USER_Primers.ipynb

{
 "cells": [
  {
   "cell_type": "code",
   "execution_count": 3,
   "metadata": {},
   "outputs": [],
   "source": [
    "import numpy as np\n",
    "import pandas as pd\n",
    "import  re\n",
    "from Bio.SeqUtils import MeltingTemp as mt\n",
    "from Bio.Seq import Seq\n",
    "from Bio.SeqUtils.MeltingTemp import make_table, DNA_NN2\n",
    "import pyperclip\n",
    "import importlib\n",
    "import DNA_tools as dt\n",
    "#https://biopython.org/docs/1.75/api/Bio.SeqUtils.MeltingTemp.html\n",
    "Breslauer= mt.DNA_NN1\n",
    "Santalucia= mt.DNA_NN4 #same as benchling\n",
    "Tm_Opt =Santalucia"
   ]
  },
  {
   "cell_type": "markdown",
   "metadata": {},
   "source": [
    "## This notebook intends to automatically design primers with overhangs for USER cloning"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 4,
   "metadata": {},
   "outputs": [
    {
     "name": "stdout",
     "output_type": "stream",
     "text": [
      "ACGGGGATU   GC%: 0.5555555555555556\n",
      "TM: 30 9\n",
      "Reverse:  AATCCCCGT\n"
     ]
    }
   ],
   "source": [
    "#Design User overhang\n",
    "HM='A'\n",
    "for i in range(7):\n",
    "    r=np.random.rand(1)\n",
    "    if r <=0.625:\n",
    "        HM+=np.random.choice(['G','C'])\n",
    "    else:\n",
    "        HM+=np.random.choice(['A','T'])\n",
    "HM+='U'\n",
    "print(HM, '  GC%:',(HM.count('C')+HM.count('G'))/len(HM))\n",
    "\n",
    "\n",
    "\n",
    "Tm=0\n",
    "for bp in HM:\n",
    "    if bp == 'A' or bp == 'T':\n",
    "        Tm+=2\n",
    "    else:\n",
    "        Tm+=4\n",
    "Rv=''\n",
    "for bp in HM[::-1]:\n",
    "    if bp == 'A':\n",
    "        Rv+='T'\n",
    "    if bp == 'T' or bp == 'U':\n",
    "        Rv+='A'\n",
    "    if bp == 'C':\n",
    "        Rv+='G'\n",
    "    if bp == 'G':\n",
    "        Rv+='C'\n",
    "        \n",
    "        \n",
    "print('TM:',Tm, len(HM))\n",
    "print('Reverse: ', Rv)"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 5,
   "metadata": {},
   "outputs": [
    {
     "name": "stdout",
     "output_type": "stream",
     "text": [
      "GC content: 0.48 \n",
      " Overhang Fwd:  TGGTCCGTGACTTGCATCGGTCATAGGGAGTTCTGTTTAATTGGTGACTC \n",
      " Overhang Rv:  GAGTCACCAATTAAACAGAACTCCCTATGACCGATGCAAGTCACGGACCA\n"
     ]
    }
   ],
   "source": [
    "#Design 50bp random overhang for yeast HR:\n",
    "\n",
    "\n",
    "continue_search = True\n",
    "while continue_search == True:\n",
    "    Overhang=''\n",
    "    for i in range(50):\n",
    "        r=np.random.rand(1)\n",
    "        \n",
    "        bp=''\n",
    "        if r<= 0.25:\n",
    "            bp='A'\n",
    "        elif r>0.25 and r<=0.5:\n",
    "            bp='T'\n",
    "        elif r>0.5 and r<=0.75:\n",
    "            bp='C'\n",
    "        else:\n",
    "            bp='G'\n",
    "        Overhang+=bp\n",
    "    \n",
    "    GC =(Overhang.count('C')+Overhang.count('G'))/len(Overhang)\n",
    "    if GC >= 0.475 and GC <= 0.525: #makes sure the overhang is balanced\n",
    "        continue_search = False\n",
    "        \n",
    "Rv_Overhang= dt.reverse_complement(Overhang)\n",
    "print('GC content:', GC, '\\n','Overhang Fwd: ', Overhang,'\\n Overhang Rv: ', Rv_Overhang)"
   ]
  },
  {
   "cell_type": "markdown",
   "metadata": {},
   "source": [
    "### Define your user overhangs:"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 6,
   "metadata": {},
   "outputs": [],
   "source": [
    "#forward seq of user overhangs:\n",
    "User_Overhangs={}\n",
    "User_Overhangs['GV1']= 'CGTGCGAu'\n",
    "User_Overhangs['GV2']= 'ATCGCGTG'#'CACGCGAu'\n",
    "User_Overhangs['U1'] = 'AGCCCTCTu'\n",
    "User_Overhangs['U2'] = 'ACACAGGCu'\n",
    "User_Overhangs['U3'] = 'ATGCCACCu'\n",
    "User_Overhangs['U4'] = 'AAGGAGCGu'\n",
    "User_Overhangs['U5'] = 'ACCGTAGCu'\n",
    "User_Overhangs['U6'] = 'ACTGCGAGu'\n",
    "User_Overhangs['U7'] = 'AGTGCAGGu'\n",
    "User_Overhangs['none'] = 'TATATATATATATA'"
   ]
  },
  {
   "cell_type": "raw",
   "metadata": {},
   "source": [
    "#part dictionary frame:\n",
    "\n",
    "part={}\n",
    "part['name']=''\n",
    "part['seq']=''\n",
    "part['need_reverse']= #if parts needs to be reversed:1 else:0\n",
    "part['Primer_up_U']='' #user site up\n",
    "part['Primer_down_U']='' #user site down\n",
    "part['Primer_up']=''\n",
    "part['Primer_up_Tm']=''\n",
    "part['Primer_down']=''\n",
    "part['Primer_down_Tm']=''"
   ]
  },
  {
   "cell_type": "markdown",
   "metadata": {},
   "source": [
    "Copy and paste sequence from benchling \n",
    "The primers will be designed taking the immediate bp at the start and end of sequence \n",
    "(bp are added to annealling sequence till Tm reaches specified Tm (or avg Tm if speficied primers))\n",
    "Always copy sequence as is (not reverse complement). \n",
    "If the seq needs to be reversed in assembly set partX['need_reverse']=1\n",
    "If you provide primers, input annealing sequence from 5'>>3'\n",
    "Specify the user sites as wanted in the final construct"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 11,
   "metadata": {},
   "outputs": [],
   "source": [
    "#Imput your parts here\n",
    "part1={}\n",
    "part1['name']='pMeLS0025(pCYC_benO::yEGFP::Cyc1t)'\n",
    "part1['seq']='gatccaggcaactttagtgctgacacataatactccataggtattttattatacaaataatgtgtttgaacttattaaaacattcttttaaggtataaacaacaggcatatatatatgtgtgcgacgacacatgatcatatggcatgcatgtgctctgtatgtatataaaactcttgttttcttcttttctctaaatattctttccttatacattaggacctttgcagcataaattactatacttctatagacacacaaacacaaatacacacactaaattaataatctgtcataaaacaatgtctaaaggtgaagaattattcactggtgttgtcccaattttggttgaattagatggtgatgttaatggtcacaaattttctgtctccggtgaaggtgaaggtgatgctacttacggtaaattgaccttaaaatttatttgtactactggtaaattgccagttccatggccaaccttagtcactactttcggttatggtgttcaatgttttgcgagatacccagatcatatgaaacaacatgactttttcaagtctgccatgccagaaggttatgttcaagaaagaactatttttttcaaagatgacggtaactacaagaccagagctgaagtcaagtttgaaggtgataccttagttaatagaatcgaattaaaaggtattgattttaaagaagatggtaacattttaggtcacaaattggaatacaactataactctcacaatgtttacatcatggctgacaaacaaaagaatggtatcaaagttaacttcaaaattagacacaacattgaagatggttctgttcaattagctgaccattatcaacaaaatactccaattggtgatggtccagtcttgttaccagacaaccattacttatccactcaatctgccttatccaaagatccaaacgaaaagagagaccacatggtcttgttagaatttgttactgctgctggtattacccatggtatggatgaattgtacaaataaatcgcgtgcattcatccgctctaaccgaaaaggaaggagttagacaacctgaagtctaggtccctatttatttttttatagttatgttagtattaagaacgttatttatatttcaaatttttcttttttttctgtacagacgcgtgtacgcatgtaacattatactgaaaaccttgcttgagaaggttttgggacgctcgaagatcgcgtcagctgaa' #sequence to clone\n",
    "part1['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "part1['Primer_up_U']='GV1' #user site up\n",
    "part1['Primer_down_U']='GV2' #user site down\n",
    "part1['Primer_up']=''\n",
    "part1['Primer_up_Tm']=''\n",
    "part1['Primer_down']=''\n",
    "part1['Primer_down_Tm']=''\n",
    "\n",
    "part2={}\n",
    "part2['name']='pMeLS0076(Rev1p::BenM(H110R, F211V, Y286N))'\n",
    "part2['seq']='gaatgcggccgctttaccaatttggtggttcagtaaaaccttcgtaggcgtagatttgtctgatggtttcgtataatgagtagatgtaggtggattcttccatgtttctaacagcaatgtagattggggtaatagcatctggatctaataatgggacgtaggacaagttgaataattgaatggattgggtagaagctggaaccaatgaaataccttcaccagcagcaaccaaacccaaggccaattggacttctctaacttcgttaatcttggtaggttccaaaccatggtcagagaagatgttcataacatgagtagagaagtttggctttggagaagatgggtacaacaagatcttttcgtcgatcaaatcgttcaagtgaacacccttatccttcatttgattcaatggatgggaagcatgaacagcaaccatcaatctttcgtttctcaacaaggttctcttgatggctggatcagaaatcttcaatctaccaaaaccagcgtcaattctaccttctttcaaagcttcggtttgagccttagtacccatttcgtataattcgattctcaagtttggatgagcttgtctgtacaagtggataattcttggcaacaaaccgaacaacaaggaaccaacaaaaccgattctaatggtcttttcaacagaggcaattctcttggtcatactgaccatttgatcaacgttgcttagcaacttgatggcgtattggtaaaagaaatgaccttctggagtagtcttaactggtctggaacctctttccaataattggatacccaattcttcttccaagttttggatttgtctggataatggtggttgagcaatacacaacttgtcggcagcctttgtaaaagattgttcttcaacaacggcaacgaagtatctcaagtgtctcaattccattgttttatgacagatcgctggatatgcctagaaatgcagtatcgatttattttgagaaaatctgtcaactcgttacttctttgatagggaaatctttagttaaatttgtttaaaattaagaaccaggtctttagactatgccaaaaggtattggtgcttaaaggttgccgtaaactctaactctatatatctctaagccaatgtttaagttgatgcgatgtgcatgatcacttcacatttttttcatgattttgttttcttccttagctacttttttccgtcttggtctgtgtaaaaaaaaaatcgcttcactgaagaaaaatataaataaaaaaaggacatcagcaagttcgtagtcaataaatgtgatgagtaatttgcgataatctaccccaactcaaagcattcaattcaaatgaaagaagagttcgtgaattttaatctggcttacagattggatcatagcttctaatatatcttcctttttattttaaagtgattaaaatatgtggatgtctgcattaaggtgaaggcagactatagtgaaaaatagtagcaaaggcttcatataatgaactacctagttgtattcaattggaatctgtgaacgcggcattaaaagaatatttgagctttttttgattgtattaacctgaagctatcacggtttagactcattaaaagaaaaatatccggttgtatcggataacaacaccgctgtagtagcacagttttttgatcatagtttcattatttttttaaatatgcttgtatacagtaatctagttttggacggattattcatgtcagtaagattttactatctactttaaaattatcgaaacagatatcaactatacatccactcattattggcatagcgaaataaaatttatgaataggaaggatataaaacgaaaattgaaatcgtttagtattatatgaaactattagtcttttcgttttcgtcgtcatacttacattcttcttttataaatattgttgtgcctaagaagagctccagcttttgttccctttagtgagggttaattgcgcgcttggcgtaatcatggtcatagctgtttcctgtgtgaaattgttatccgct'\n",
    "part2['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "part2['Primer_up_U']='GV1' #user site up\n",
    "part2['Primer_down_U']='GV2' #user site down\n",
    "part2['Primer_up']=''\n",
    "part2['Primer_up_Tm']=''\n",
    "part2['Primer_down']=''\n",
    "part2['Primer_down_Tm']=''\n",
    "\n",
    "part2B={}\n",
    "part2B['name']='pMeLS0076(Rev1p::BenM_MP17_D08)'\n",
    "part2B['seq']='Ttaccaatttggtggttcagtaaaaccttcgtaggcgtagatttgtctgatggtttcgtataatgagtagatgtaggtggattcttccatgtttctaacagcaatgtagattggggtaatagcatctggatctaataatgggacgtaggacaagttgaataattgaatggattgggtagaagctggaaccaatgaaataccttcaccagcagcaaccaaacccaaggccaattggacttctctaacttcgttaatcttggtaggttccaaaccatggtcagagaagatgttcataacatgagtagagaagtttggctttggagaagatgggtacaacaagatcttttcgtcgatcaaatcgttcaagtgaacacccttatccttcatttgattcaatggatgggaagcatgaacagcaaccatcaatctttcgtttctcaacaaggttctcttgatggctggatcagaaatcttcaatctaccaaaaccagcgtcaattctaccttctttcaaagcttcggtttgagccttagtacccatttcgtataattcgattctcaagtttggatgagcttgtctgtacaagtggataattcttggcaacaaaccgaacaacaaggaaccaacaaaaccgattctaatggtcttttcaacagaggcaattctcttggtcatactgaccatttgatcaacgttgcttagcaacttgatggcgtattggtaaaagaaatgaccttctggagtagtcttaactggtctggaacctctttccaataattggatacccaattcttcttccaagttttggatttgtctggataatggtggttgagcaatacacaacttgtcggcagcctttgtaaaagattgttcttcaacaacggcaacgaagtatctcaagtgtctcaattccattgttttatgacagatcgctggatatgcctagaaatgcagtatcgatttattttgagaaaatctgtcaactcgttacttctttgatagggaaatctttagttaaatttgtttaaaattaagaaccaggtctttagactatgccaaaaggtattggtgcttaaaggttgccgtaaactctaactctatatatctctaagccaatgtttaagttgatgcgatgtgcatgatcacttcacatttttttcatgattttgttttcttccttagctacttttttccgtcttggtctgtgtaaaaaaaaaatcgcttcactgaagaaaaatataaataaaaaaaggacatcagcaagttcgtagtcaataaatgtgatgagtaatttgcgataatctaccccaactcaaagcattcaattcaaatgaaagaagagttcgtgaattttaatctggcttacagattggatcatagcttctaatatatcttcctttttattttaaagtgattaaaatatgtggatgtctgcattaaggtgaaggcagactatagtgaaaaatagtagcaaaggcttcatataatgaactacctagttgtattcaattggaatctgtgaacgcggcattaaaagaatatttgagctttttttgattgtattaacctgaagctatcacggtttagactcattaaaagaaaaatatccggttgtatcggataacaacaccgctgtagtagcacagttttttgatcatagtttcattatttttttaaatatgcttgtatacagtaatctagttttggacggattattcatgtcagtaagattttactatctactttaaaattatcgaaacagatatcaactatacatccactcattattggcatagcgaaataaaatttatgaataggaaggatataaaacgaaaattgaaatcgtttagtattatatgaaactattagtcttttcgttttcgtcgtcatacttacattcttcttttataaatattgttgtgcctaagaa'\n",
    "part2B['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "part2B['Primer_up_U']='GV1' #user site up\n",
    "part2B['Primer_down_U']='GV2' #user site down\n",
    "part2B['Primer_up']=''\n",
    "part2B['Primer_up_Tm']=''\n",
    "part2B['Primer_down']=''\n",
    "part2B['Primer_down_Tm']=''\n",
    "\n",
    "\n",
    "part3={}\n",
    "part3['name']='pTDH2'\n",
    "part3['seq']='tttgttttgtttgtttgtgtgatgaatttaatttgaaactaagttcttggtttttttaacatttaaacaaacaataataaaatgaatagacacacaagaaaaaaactatcttgaatgagtgcaacctttatatattcttccaaacctacaaaaaaattgcaggtctgcgatcactgggtactatcaaaacacatattagcatacgtatgtgttcgtacgtgcggtccgcaaggtctttgtgacagccatgacatgattggattgaatagttcacttcagctgtcaccaccagttttaggccctaacacatcagtcgagttccactaggctttgttagccgtagctgcgttgcaacaactgcgcaaaacgcgcgacttcctgtaaatgccgcagatcacgtggtaaagaaaagaaacaacaggccgacccagtttcgttgtttcttttcattatttctggatgctcttcacaacgtatttcatgacatacttattgaattagtggaagcccaaaacatcattaccctggaaaacttaattgacgtaaattaacttcagttgtcttatgttttttgcatccttaactttagccgcgcagggataatcatatatgtcgaaagagcaactaaacaatacttaaaaacgtgtttgaacttttacgaaagggtaacgacaaataatcattcactggaaaaaccaataaaccataagtaaatgaaaaactgtttcaaccgatgttggtactttccatgtcttaaaagagtacaatttataaattaatgaaaacacagaagtatttagatcggctcaaatgtttttggacattaaaagatcttgaaactgagtaagatgctcagaatacccgtcaagataagagtataatgtagagtaatataccaagtattcagcatattctcctcttcttttgtataaatcacggaagggatgatttataagaaaaatgaatactattacacttcatttaccaccctctgatctagat'\n",
    "part3['need_reverse']=1 #if parts needs to be reversed:1 else:0\n",
    "part3['Primer_up_U']='GV1' #user site up\n",
    "part3['Primer_down_U']='U1' #user site down\n",
    "part3['Primer_up']=''\n",
    "part3['Primer_up_Tm']=''\n",
    "part3['Primer_down']='aaatctagatcagagggtggtaaat'\n",
    "part3['Primer_down_Tm']=''\n",
    "\n",
    "part4={}\n",
    "part4['name']='pCFB1237_ScTkl1'\n",
    "part4['seq']='tcagaaagcttttttcaaaggagaaattagcttgtcacccttatagaatgcaatggtcttttgagctctttcagcaacaccttctggggtgaaaccgaagaacttgaagacttctggtgccttaccggaggcaccaaatctgtcaataccgaaggattgatgagcgtatttgccccaacatgtggtagccaaaacttcaacagacatgattggaacgttgtctggtaagactgatagtctgtattctaggggttgtttgtcaaaagtgaagaaatctggtagagaaacaacacgagccttgatgttctttgcggccaaagtcttagcagcttcaacactcaaagacacttcggaaccagtagccactaaaataatatctgggttagcaacatcttgtagtacgtaaccacccttagaagcgctttcaatagagctaccttccaattgtggcaagttttgtctggacaaagcaatgatacttggagtatgcttggattctaaagagttcttgtaggcggcagaaacttcgttaccatcagctggtctccaaacttgaatgtttggtagggatctgaagtgtgctaaagtttcaataggttgatgtgttggaccatcttcaccgacaccgatagagtcatgtgtagcaacccaaataactgggtggccagacaaagcggacaatctaacggcaccagcagcataagaaacgaagttcaagaaagtaccaccgtatggtttgtagttggcaccgaaagctgaaataccgttcattatggcacccatagcgtgttctctaataccgtacctaatgtatctaccagagtagttacctgaaccggaagaaggaggttggaagtcaagggcttccttccatctggtcaagttagaaggtgttaaatcggcagaaccaccaatcaactctggcaattgattgtaaacatcctcaagaacagtttctgataattttctagtggccacggcagagtccttggcggtgtaagttggcaacttagattcccaatttgcgggtagttggccgctcaatcttctagccaattcagcacctaattctgggaatttcttttggtattcgctgaacaacttgttccacttgttgttggcttcgacacctggctttaaaattgtcttttggtagtggtcgtaaacttcttgtggaacaacaaaggacttgtctgggttgaaaccgaatttgctctttagttgtttaacatcatctgctttcaatggggcaccgtgcacagagtgagagccggcatgcaaggaaccgtaaccaatggttgtggtcattttgatcaaagttggtttgtccttggataacttagcttgagcaatagccttggcaataccggctagatcttcgttaccattttctacgtacaaaacttcccaaccgtaggcttcgtatctcttagcaacatcttcatcgaatgagatactggtagcaccatcgatagtgatcttgttgtcatcgtagatggcaatcaagttacccaatttcaaatgaccagccaaggaggaagcttctgaagaaataccttcttgcaaacaaccgtcacccaagaaaacataggtgtagttgtcagacaaggtaaagcccggcttgttgtaagtggcagccaggttagcttgagccatggccataccaacagcgttggagataccttgacctaatggaccggtagtaacttcaacacctggcaactcaaattcaggatgacctggtgttctggaacccaactgtctgaactgtttcaagtcttcaatagacagatcgtaaccagtcaaatgtagcatagaatacaacaaagcgaccgcgtgaccgttagacaagacaaatctatctctgttgatccagtctgggttggttgggttcatgcgcatttgactccatagaacgtgtgcagctggtgccatacccaatggagcacctgggtgacctgagttggccttggatacggtgtccacagccaaaattcttatggtggagacggctagcttatcaatgtcagtgaattgagtcattgt'\n",
    "part4['need_reverse']= 1 #if parts needs to be reversed:1 else:0\n",
    "part4['Primer_up_U']='U1' #user site up\n",
    "part4['Primer_down_U']='U2' #user site down\n",
    "part4['Primer_up']=''\n",
    "part4['Primer_up_Tm']=''\n",
    "part4['Primer_down']=''\n",
    "part4['Primer_down_Tm']=''\n",
    "part5={}\n",
    "part5['name']='ANTE113_Tdh3bidir'\n",
    "part5['seq']='TCTGCAGGTAGGGAAAGATATGAGCTATACAGCGGAATTTCCATATCACTCAGATTTTGTTATCTAATTTTTTCCTTCCCACGTCCGCGGGAATCTGTGTATATTACTGCATCTAGATATATGTTATCTTATCTTGGCGCGTACATTTAATTTTCAACGTATTCTATAAGAAATTGCGGGAGTTTTTTTCATGTAGATGATACTGACTGCACGCAAATATAGGCATGATTTATAGGCATGATTTGATGGCTGTACCGATAGGAACGCTAAGAGTAACTTCAGAATCGTTATCCTGGCGGAAAAAATTCATTTGTAAACTTTAAAAAAAAAAGCCAATATCCCCAAAATTATTAAGAGCGCCTCCATTATTAACTAAAATTTCACTCAGCATCCACAATGTATCAGGTATCTACTACAGATATTACATGTGGCGAAAAAGACAAGAACAATGCAATAGCGCATCAAGAAAAAACACAAAGCTTTCAATCAATGAATCGAAAATGTCATTAAAATAGTATATAAATTGAAACTAAGTCATAAAGCTATAAAAAGAAAATTTATTTAAATGCAAGATTTAAAGTAAATTCAC'\n",
    "part5['need_reverse']= 1 #if parts needs to be reversed:1 else:0\n",
    "part5['Primer_up_U']='U2' #user site up\n",
    "part5['Primer_down_U']='GV2' #user site down\n",
    "part5['Primer_up']=''\n",
    "part5['Primer_up_Tm']=''\n",
    "part5['Primer_down']=''\n",
    "part5['Primer_down_Tm']=''\n",
    "\n",
    "\n",
    "User1=[part1]\n",
    "User2=[part2]\n",
    "User2B=[part2B] #pCCM010\n",
    "User3=[part3, part4,part5]"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 12,
   "metadata": {},
   "outputs": [],
   "source": [
    "part6={}\n",
    "part6['name']='FBA1p'\n",
    "part6['seq']='tttgaatatgtattacttggttatggttatatatgacaaaagaaaaagaagaacagaagaataacgcaaggaagaacaataactgaaattgatagagaagtattatgtctttgtctttttataataaatcaagtgcagaaatccgttagacaacatgagggataaaatttaacgtgggcgaagaagaaggaaaaaagtttttgtgagggcgtaattgaagcgatctgttgattgtagattttttttttttgaggagtcaaagtcagaagagaacagacaaatggtattaaccatccaatacttttttggagcaacgctaagctcatgcttttccattggttacgtgctcagttgttagatatggaaagagaggatgctcacggcagcgtgactccaattgagcccgaaagagaggatgccacgttttcccgacggctgctagaatggaaaaaggaaaaatagaagaatcccattcctatcattatttacgtaatgacccacacatttttgagattttcaactattacgtattacgataatcctgctgtcattatcattattatctatatcgacgtatgcaacgtatgtgaagccaagtaggcaattatttagtactgtcagtattgttattcatttcttagggcttgcgtcagcttgatgaggatttgtgatagtagccgttggaggcatgaagggcattggaaacatggggaaaggaggtataccgaaaggtaacggtactggtggcactggcgtaccgccgttatttacagaagttggaaggctggtattgttgttcaagccagcggtgccagttggatccatcaacttcattttcttagtttctggctggttgccatctttagagcttccgtgtagttcctcttgtttcttcaaaaacgtttcgacctccttttccttgtcctggtcgggtactaaagaatcgagaaggatatcgcgggtttcgcaattggggcatacaaagtcgctctctaccagt'\n",
    "part6['need_reverse']=1 #if parts needs to be reversed:1 else:0\n",
    "part6['Primer_up_U']='GV1' #user site up\n",
    "part6['Primer_down_U']='U1' #user site down\n",
    "part6['Primer_up']='tttgaatatgtattacttggttatg'\n",
    "part6['Primer_up_Tm']=''\n",
    "part6['Primer_down']='actggtagagagcgacttt'\n",
    "part6['Primer_down_Tm']=''\n",
    "\n",
    "part7={}\n",
    "part7['name']='pCFB1239_PaAroZ'\n",
    "part7['seq']='tcacaaagcagctgacaaagacaacattctgataggttgttgtgtagcttgttgttgagttggagagtcaacggtgatcttcatatctctaaccaatttagcccaagaaacagcacctcttctagctaattcttctggaacaccaaaaccggtatcggacattcttctgttgaataattctaaggaaacccaaccttcaaaacccaaaccgttgaaaaaagcccaagcaatttcttttactggcaagtaagcacctctatccttttcaccgtagaataatctgcagtttctagaccaagacattctagctggttgttctggatcatagaatctgtgacctggaaccaatggtttcttcaacttttcggcatcaacaacttgaacgtagaaaaccttggaaacgtcaactctttcaaccaatctggcaatggactttctgatagcttcttcagcatttggagttctaccagaagcaacagttggatcagcataaactctaccggcaatgttaaaagtgtccaaacaaacaccgaagtttggtctgttaactctttgaacaacttcccaagatctttcccaagtatcaactctagtggaccaacacaaagcttcgtaaacgaatctaattggtgggttagcttgtaaacccatatcagctacttcttgcaagtcggagacaatcaaagagatatcttcagtaacttcttcggctggcaagaagttggctggaatttgaataatatcggtatccaattcatgggccaattcgatccagaattccaattgttccaatcttctttcgtgttcttctctatccaacaaaccatcgtattgagagaatggttgcaagcaaacgatttcgatgtttctgacttggcacattctcaaaatttgtctagcagcagacaattgagcagctggtgatggaccacatggagatggagtttcaccagacaatctataggcgacatcagccaaatcttcatggaataattccaaaccttggtaaccgtactttctagccatatccaacttagtagtgaaagaatgaccggcataacatctacccaaagacatagaggtaatggccaacttggatggcattgt'\n",
    "part7['need_reverse']= 1 #if parts needs to be reversed:1 else:0\n",
    "part7['Primer_up_U']='U1' #user site up\n",
    "part7['Primer_down_U']='U2' #user site down\n",
    "part7['Primer_up']=''\n",
    "part7['Primer_up_Tm']=''\n",
    "part7['Primer_down']=''\n",
    "part7['Primer_down_Tm']=''\n",
    "\n",
    "part8={}\n",
    "part8['name']='His5t'\n",
    "part8['seq']='atagattaatttaaacagtatatgtacagttttatatatatatatatatatatatacatatataaagaaacctgtgcgttttttgtattttcaaatacatttagttttggcgccacttctataaaaggtctcatgatattgttac'\n",
    "part8['need_reverse']= 0  #if parts needs to be reversed:1 else:0\n",
    "part8['Primer_up_U']='U2' #user site up\n",
    "part8['Primer_down_U']='GV2' #user site down\n",
    "part8['Primer_up']='atagattaatttaaacagtatatgtacagttt'\n",
    "part8['Primer_up_Tm']=''\n",
    "part8['Primer_down']='gtaacaatatcatgagaccttttata'\n",
    "part8['Primer_down_Tm']=''\n",
    "\n",
    "part9={}\n",
    "part9['name']='TPI1p'\n",
    "part9['seq']='ttttagtttatgtatgtgttttttgtagttatagatttaagcaagaaaagaatacaaacaaaaaattgaaaaagattgatttagaattaaaaagaaaaatatttacgtaagaagggaaaatagtaaatgttgcaagttcactaaactcctaaattatgctgccctttatattccctgttacagcagccgagccaaaggtatataggctcctttgcattagcatgcgtaacaaaccacctgtcagtttcaaccgaggtggtatccgagagaattgtgtgattgctttaattaatttcggagaatctcacatgccactgaagattaaaaactggatgccagaaaaggggtgtccaggtgtaacatcaatagaggaagctgaaaagtcttagaacgggtaatcttccaccaacctgatgggttcctagatataatctcgaagggaataagtagggtgataccgcagaagtgtctgaatgtattaaggtcctcacagtttaaatcccgctcacactaacgtaggattattataactcaaaaaaatggcattattctaagtaagttaaatatccgtaatctttaaacactatgtagttaggtctccctcacaatcagtccatttgggtagcacggtcctcaggacgtatctattgatggattcgtccagttccatcaccattacgctcccgttaggaacattggtaaacgattcaaactcttcgtatgtccatctaaaccatttcatcaggaatactctggaataaataccatgtgtaactaggacaacaacatctctgggtcttctctcttgcctatcatggaagtgcctgaataaagtctcttggaaactggcgactctgtcatatacatctgccgcactttctccatgagggaatctgaagaagaaatgaccatacgtagatctcttcttcataacgtcctgcatgctattgattttttggaaattaccaaaatcttgttcccttattcttggctcatcctt'\n",
    "part9['need_reverse']=1 #if parts needs to be reversed:1 else:0\n",
    "part9['Primer_up_U']='GV1' #user site up\n",
    "part9['Primer_down_U']='U1' #user site down\n",
    "part9['Primer_up']=''\n",
    "part9['Primer_up_Tm']=''\n",
    "part9['Primer_down']=''\n",
    "part9['Primer_down_Tm']=''\n",
    "\n",
    "part10={}\n",
    "part10['name']='pCfB2193_NatMX'\n",
    "part10['seq']='accatgggtaccactcttgacgacacggcttaccggtaccgcaccagtgtcccgggggacgccgaggccatcgaggcactggatgggtccttcaccaccgacaccgttttccgcgtcaccgccaccggggacggcttcaccctgcgggaggtgccggtggacccgcccctgaccaaggtgttccccgacgacgaatcggacgacgaatcggacgacggggaggacggcgacccggactcccggacgttcgtcgcgtacggggacgacggcgacctggcgggcttcgtggtcgtctcgtactccggctggaaccgccggctgaccgtcgaggacatcgaggtcgccccggagcaccgggggcacggggtcgggcgcgcgttgatggggctcgcgacggagttcgcccgcgagcggggcgccgggcacctctggctggaggtcaccaacgtcaacgcaccggcgatccacgcgtaccggcggatggggttcaccctctgcggcctggacaccgccctgtacgacggcaccgcctcggacggcgagcaggcgctctacatgagcatgccctgcccctaa'\n",
    "part10['need_reverse']= 0#if parts needs to be reversed:1 else:0\n",
    "part10['Primer_up_U']='U1' #user site up\n",
    "part10['Primer_down_U']='U2' #user site down\n",
    "part10['Primer_up']=''\n",
    "part10['Primer_up_Tm']=''\n",
    "part10['Primer_down']=''\n",
    "part10['Primer_down_Tm']=''\n",
    "\n",
    "part11={}\n",
    "part11['name']='LSC2t'\n",
    "part11['seq']='aaaattaaaaaaaaaaaaaagaaattttttccaaaagagtatatttatatgtatgtatacatgtagggaaaataagaaactttattaatagtaaaaaagcatatatactttattattaactcttttgtttttctcgagaagc'\n",
    "part11['need_reverse']=1 #if parts needs to be reversed:1 else:0\n",
    "part11['Primer_up_U']='U2' #user site up\n",
    "part11['Primer_down_U']='GV2' #user site down\n",
    "part11['Primer_up']='aaaattaaaaaaaaaaaaaagaaattttttcc'\n",
    "part11['Primer_up_Tm']=''\n",
    "part11['Primer_down']='gcttctcgagaaaaacaaaagagtta'\n",
    "part11['Primer_down_Tm']=''\n",
    "\n",
    "part12={}\n",
    "part12['name']='pCYC1'\n",
    "part12['seq']='cagcattttcaaaggtgtgttcttcgtcagacatgttttagtgtgtgaatgaaataggtgtatgttttctttttgctagacaataattaggaacaaggtaagggaactaaagtgtagaataagattaaaaaagaagaacaagttgaaaaggcaagttgaaatttcaagaaaaaagtcaattgaagtacagtaaattgacctgaatatatctgagttccgacaacaatgagtttaccaaagagaacaatggaataggaaactttgaacgaagaaaggaaagcaggaaaggaaaaaatttttaggctcgagaacaatagggcgaaaaaacaggcaacgaacgaacaatggaaaaacgaaaaaaaaaaaaaaaaacacagaaaagaatgcagaaagatgtcaactgaaaaaaaaaaaggtgaacacaggaaaaaaaataaaaaaaaaaaaaaaaaaaggaggacgaaacaaaaaagtgaaaaaaaatgaaaatttttttggaaaaccaagaaatgaattatatttccgtgtgagacgacatcgtcgaatatgattcagggtaacagtattgatgtaatcaatttcctacctgaatctaaaattcccgggagcaagatcaagatgttttcaccgatctttccggtctctttggccggggtttacggacgatggcagaagaccaaagcgccagttcatttggcgagcgttggttggtggatcaagcccacgcgtaggcaatcctcgagcagatccgccaggcgtgtatatatagcgtggatggccaggcaactttagtgctgacacatacaggcatatatatatgtgtgcgacgacacatgatcatatggcatgcatgtgctctgtatgtatataaaactcttgttttcttcttttctctaaatattctttccttatacattaggacctttgcagcataaattactatacttctatagacacacaaacacaaatacacacactaaattaata'\n",
    "part12['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "part12['Primer_up_U']='GV1' #user site up\n",
    "part12['Primer_down_U']='U1' #user site down\n",
    "part12['Primer_up']='cagcattttcaaaggtgt'\n",
    "part12['Primer_up_Tm']=''\n",
    "part12['Primer_down']=''\n",
    "part12['Primer_down_Tm']=''\n",
    "\n",
    "part13={}\n",
    "part13['name']='pCFB1239CaCatA'\n",
    "part13['seq']='acaatgtcccaagctttcaccgaatctgttaagacttctttgggtccaaatgctactccaagagctaaaaagttgattgcctctttggttcaacacgttcatgatttcgctagagaaaaccatttgactaccgaagattggttgtggggtgttgatttcattaacagaattggtcaaatgtccgactccagaagaaacgaaggtattttggtttgcgatatcatcggtttggaaaccttggttgatgctttgactaacgaatccgaacaatctaaccatacctcctctgctattttgggtcctttttacttgccagattctccagtttatccaaacggtggttctatcgttcaaaaggctattccaactgatgttaagtgcttcgttagaggtaaggttactgatactgaaggtaaaccattgggtggtgctcaattggaagtttggcaatgtaattctgctggtttctactctcaacaagctgatcatgatggtccagaattcaatttgagaggtactttcattaccgacgacgaaggtaattactccttcgaatgtttaagaccaacctcctatccaattccatacgatggtcctgctggtgatttgttgaaaatcatggatagacatccaaacagaccatcccatattcattggagagtttctcatccaggttaccatactttgatcacccaaatctatgatgctgaatgtccatacaccaacaacgattctgtttacgctgttaaggatgacatcatcgttcacttcgaaaaggttgataacaaggataaggatttggtcggtaaggtcgaatacaagttggattacgatatttccttggccaccgaatcctctattcaagaagctagagctgctgctaaagctagacaagatgctgaaatcaagttgtaa'\n",
    "part13['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "part13['Primer_up_U']='U1' #user site up\n",
    "part13['Primer_down_U']='U2' #user site down\n",
    "part13['Primer_up']=''\n",
    "part13['Primer_up_Tm']=''\n",
    "part13['Primer_down']=''\n",
    "part13['Primer_down_Tm']=''\n",
    "\n",
    "part14={}\n",
    "part14['name']='CPS1t'\n",
    "part14['seq']='gcgcaatgattgaatagtcaaagattttttttttttaattttttttttttaattttttttttttttcatagaactttttatttaaataaatcacgtctatatatgtatcagtataacgtaaaaaaaaaaacaccgtcagttaaacaaaacataaataaaaaaaaaaagaagtgtcaaatcaagtgtcaaatc'\n",
    "part14['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "part14['Primer_up_U']='U2' #user site up\n",
    "part14['Primer_down_U']='GV2' #user site down\n",
    "part14['Primer_up']='gcgcaatgattgaatagtcaaa'\n",
    "part14['Primer_up_Tm']=''\n",
    "part14['Primer_down']='gatttgacacttgatttgacacttcttt'\n",
    "part14['Primer_down_Tm']=''\n",
    "\n",
    "User4=[part6, part7,part8]\n",
    "User5=[part9, part10,part11]\n",
    "User6=[part12, part13,part14]"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 13,
   "metadata": {},
   "outputs": [],
   "source": [
    "part15={}\n",
    "part15['name']='pCfB1241_TDH3p:KpAroY.B'\n",
    "part15['seq']='tcattcaatttcttgagcgaattgttcagcagttctcaaaccgttccatcttctagctttgtgataatccaaaccgaattggtccaaaactctggtaactatatggttggtgatgtcatcaacggtttctggatgattgtaataagctggcattggtggaaccatagctacacccattctagacaaggccaacatgttttccaaatggatagtagacaatggcatttctcttgggaccaagaccaactttctaccttctttcaaaacaacatcagcagctctaccaaccaaaccttcagcataaccagctctaataccagccaaggttttcatagagcatggaataacgatcataccatcagttctgaaagaaccagaagaaatagtagcagcttgatcagctggagaatgagaaaaatcagccaaagcagcaacttctctagcagtccatggagtttccaattcaatggtggtcttagcccatttagacataaccaaatgggtttcaacttctggcatatctctcaaagcttgcaacaaagcaacacccaatggagcacctgtagcaccagtcataccgatgatcaacttcattgttttacctgcactacctgcacttttgtttgtttatgtgtgtttattcgaaactaagttcttggtgttttaaaactaaaaaaaagactaactataaaagtagaatttaagaagtttaagaaatagatttacagaattacaatcaatacctaccgtctttatatacttattagtcaagtaggggaataatttcagggaactggtttcaaccttttttttcagctttttccaaatcagagagagcagaaggtaatagaaggtgtaagaaaatgagatagatacatgcgtgggtcaattgccttgtgtcatcatttactccaggcaggttgcatcactccattgaggttgtgcccgttttttgcctgtttgtgcccctgttctctgtagttgcgctaagagaatggacctatgaactgatggttggtgaagaaaacaatattttggtgctgggattctttttttttctggatgccagcttaaaaagcgggctccattatatttagtggatgccaggaataaactgttcacccagacacctacgatgttatatattctgtgtaacccgccccctattttgggcatgtacgggttacagcagaattaaaaggctaattttttgactaaataaagttaggaaaatcactactattaattatttacgtattctttgaaatggcagtattgataatgataaactcgaactgaaaaagcgtgttttttatagc'\n",
    "#'caacggaatgcgtgcgattcattcaatttcttgagcgaattgttcagcagttctcaaaccgttccatcttctagctttgtgataatccaaaccgaattggtccaaaactctggtaactatatggttggtgatgtcatcaacggtttctggatgattgtaataagctggcattggtggaaccatagctacacccattctagacaaggccaacatgttttccaaatggatagtagacaatggcatttctcttgggaccaagaccaactttctaccttctttcaaaacaacatcagcagctctaccaaccaaaccttcagcataaccagctctaataccagccaaggttttcatagagcatggaataacgatcataccatcagttctgaaagaaccagaagaaatagtagcagcttgatcagctggagaatgagaaaaatcagccaaagcagcaacttctctagcagtccatggagtttccaattcaatggtggtcttagcccatttagacataaccaaatgggtttcaacttctggcatatctctcaaagcttgcaacaaagcaacacccaatggagcacctgtagcaccagtcataccgatgatcaacttcattgttttacctgcactacctgcacttttgtttgtttatgtgtgtttattcgaaactaagttcttggtgttttaaaactaaaaaaaagactaactataaaagtagaatttaagaagtttaagaaatagatttacagaattacaatcaatacctaccgtctttatatacttattagtcaagtaggggaataatttcagggaactggtttcaaccttttttttcagctttttccaaatcagagagagcagaaggtaatagaaggtgtaagaaaatgagatagatacatgcgtgggtcaattgccttgtgtcatcatttactccaggcaggttgcatcactccattgaggttgtgcccgttttttgcctgtttgtgcccctgttctctgtagttgcgctaagagaatggacctatgaactgatggttggtgaagaaaacaatattttggtgctgggattctttttttttctggatgccagcttaaaaagcgggctccattatatttagtggatgccaggaataaactgttcacccagacacctacgatgttatatattctgtgtaacccgccccctattttgggcatgtacgggttacagcagaattaaaaggctaattttttgactaaataaagttaggaaaatcactactattaattatttacgtattctttgaaatggcagtattgataatgataaactcgaactgaaaaagcgtgttttttatagc'\n",
    "part15['need_reverse']=1#if parts needs to be reversed:1 else:0\n",
    "part15['Primer_up_U']='GV1' #user site up\n",
    "part15['Primer_down_U']='U2' #user site down\n",
    "part15['Primer_up']=''\n",
    "part15['Primer_up_Tm']=''\n",
    "part15['Primer_down']=''\n",
    "part15['Primer_down_Tm']=''\n",
    "\n",
    "\n",
    "part16={}\n",
    "part16['name']='VPS13t'\n",
    "part16['seq']='gcgcgctgcggatatttctaaggcatggtcgtgcggagctacaataatacgattgaattatagctacatagtgtacaaaagcgggtatatactttcatatgtga'\n",
    "part16['need_reverse']=1#if parts needs to be reversed:1 else:0\n",
    "part16['Primer_up_U']='U2' #user site up\n",
    "part16['Primer_down_U']='GV2' #user site down\n",
    "part16['Primer_up']='' #we dont provide Curran's primer because their TM is very high for VPS13t\n",
    "part16['Primer_up_Tm']=''\n",
    "part16['Primer_down']='' #we dont provide Curran's primer because their TM is very high for VPS13t\n",
    "part16['Primer_down_Tm']=''\n",
    "\n",
    "User7=[part15, part16]\n"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 14,
   "metadata": {},
   "outputs": [],
   "source": [
    "# this user assembly was redesigned to accomodate a gBlock for AroyB P146T\n",
    "\n",
    "\n",
    "part1={}\n",
    "part1['name']='pCfB1241_TDH3p'\n",
    "part1['seq']='tttgtttgtttatgtgtgtttattcgaaactaagttcttggtgttttaaaactaaaaaaaagactaactataaaagtagaatttaagaagtttaagaaatagatttacagaattacaatcaatacctaccgtctttatatacttattagtcaagtaggggaataatttcagggaactggtttcaaccttttttttcagctttttccaaatcagagagagcagaaggtaatagaaggtgtaagaaaatgagatagatacatgcgtgggtcaattgccttgtgtcatcatttactccaggcaggttgcatcactccattgaggttgtgcccgttttttgcctgtttgtgcccctgttctctgtagttgcgctaagagaatggacctatgaactgatggttggtgaagaaaacaatattttggtgctgggattctttttttttctggatgccagcttaaaaagcgggctccattatatttagtggatgccaggaataaactgttcacccagacacctacgatgttatatattctgtgtaacccgccccctattttgggcatgtacgggttacagcagaattaaaaggctaattttttgactaaataaagttaggaaaatcactactattaattatttacgtattctttgaaatggcagtattgataatgataaactcgaactgaaaaagcgtgttttttatagc'\n",
    "#'caacggaatgcgtgcgattcattcaatttcttgagcgaattgttcagcagttctcaaaccgttccatcttctagctttgtgataatccaaaccgaattggtccaaaactctggtaactatatggttggtgatgtcatcaacggtttctggatgattgtaataagctggcattggtggaaccatagctacacccattctagacaaggccaacatgttttccaaatggatagtagacaatggcatttctcttgggaccaagaccaactttctaccttctttcaaaacaacatcagcagctctaccaaccaaaccttcagcataaccagctctaataccagccaaggttttcatagagcatggaataacgatcataccatcagttctgaaagaaccagaagaaatagtagcagcttgatcagctggagaatgagaaaaatcagccaaagcagcaacttctctagcagtccatggagtttccaattcaatggtggtcttagcccatttagacataaccaaatgggtttcaacttctggcatatctctcaaagcttgcaacaaagcaacacccaatggagcacctgtagcaccagtcataccgatgatcaacttcattgttttacctgcactacctgcacttttgtttgtttatgtgtgtttattcgaaactaagttcttggtgttttaaaactaaaaaaaagactaactataaaagtagaatttaagaagtttaagaaatagatttacagaattacaatcaatacctaccgtctttatatacttattagtcaagtaggggaataatttcagggaactggtttcaaccttttttttcagctttttccaaatcagagagagcagaaggtaatagaaggtgtaagaaaatgagatagatacatgcgtgggtcaattgccttgtgtcatcatttactccaggcaggttgcatcactccattgaggttgtgcccgttttttgcctgtttgtgcccctgttctctgtagttgcgctaagagaatggacctatgaactgatggttggtgaagaaaacaatattttggtgctgggattctttttttttctggatgccagcttaaaaagcgggctccattatatttagtggatgccaggaataaactgttcacccagacacctacgatgttatatattctgtgtaacccgccccctattttgggcatgtacgggttacagcagaattaaaaggctaattttttgactaaataaagttaggaaaatcactactattaattatttacgtattctttgaaatggcagtattgataatgataaactcgaactgaaaaagcgtgttttttatagc'\n",
    "part1['need_reverse']=1#if parts needs to be reversed:1 else:0\n",
    "part1['Primer_up_U']='GV1' #user site up\n",
    "part1['Primer_down_U']='U7' #user site down\n",
    "part1['Primer_up']=''\n",
    "part1['Primer_up_Tm']=''\n",
    "part1['Primer_down']=''\n",
    "part1['Primer_down_Tm']=''\n",
    "\n",
    "part2={}\n",
    "part2['name']='KpAroY.B-P146T.gBlock'\n",
    "part2['seq']='aaacaatgaagttgatcatcggtatgactggtgctacaggtgctccattgggtgttgctttgttgcaagctttgagagatatgccagaagttgaaacccatttggttatgtctaaatgggctaagaccaccattgaattggaaactccatggactgctagagaagttgctgctttggctgatttttctcattctccagctgatcaagctgctactatttcttctggttctttcagaactgatggtatgatcgttattccatgctctatgaaaaccttggctggtattagagctggttatgctgaaggtttggttggtagagctgctgatgttgttttgaaagaaggtagaaagttggtcttggtcccaagagaaatgccattgtctactatccatttggaaaacatgttggccttgtctagaatgggtgtagctatggttAcaccaatgccagcttattacaatcatccagaaaccgttgatgacatcaccaaccatatagttaccagagttttggaccaattcggtttggattatcacaaagctagaagatggaacggtttgagaactgctgaacaattcgctcaagaaattgaatga'\n",
    "#'caacggaatgcgtgcgattcattcaatttcttgagcgaattgttcagcagttctcaaaccgttccatcttctagctttgtgataatccaaaccgaattggtccaaaactctggtaactatatggttggtgatgtcatcaacggtttctggatgattgtaataagctggcattggtggaaccatagctacacccattctagacaaggccaacatgttttccaaatggatagtagacaatggcatttctcttgggaccaagaccaactttctaccttctttcaaaacaacatcagcagctctaccaaccaaaccttcagcataaccagctctaataccagccaaggttttcatagagcatggaataacgatcataccatcagttctgaaagaaccagaagaaatagtagcagcttgatcagctggagaatgagaaaaatcagccaaagcagcaacttctctagcagtccatggagtttccaattcaatggtggtcttagcccatttagacataaccaaatgggtttcaacttctggcatatctctcaaagcttgcaacaaagcaacacccaatggagcacctgtagcaccagtcataccgatgatcaacttcattgttttacctgcactacctgcacttttgtttgtttatgtgtgtttattcgaaactaagttcttggtgttttaaaactaaaaaaaagactaactataaaagtagaatttaagaagtttaagaaatagatttacagaattacaatcaatacctaccgtctttatatacttattagtcaagtaggggaataatttcagggaactggtttcaaccttttttttcagctttttccaaatcagagagagcagaaggtaatagaaggtgtaagaaaatgagatagatacatgcgtgggtcaattgccttgtgtcatcatttactccaggcaggttgcatcactccattgaggttgtgcccgttttttgcctgtttgtgcccctgttctctgtagttgcgctaagagaatggacctatgaactgatggttggtgaagaaaacaatattttggtgctgggattctttttttttctggatgccagcttaaaaagcgggctccattatatttagtggatgccaggaataaactgttcacccagacacctacgatgttatatattctgtgtaacccgccccctattttgggcatgtacgggttacagcagaattaaaaggctaattttttgactaaataaagttaggaaaatcactactattaattatttacgtattctttgaaatggcagtattgataatgataaactcgaactgaaaaagcgtgttttttatagc'\n",
    "part2['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "part2['Primer_up_U']='U7' #user site up\n",
    "part2['Primer_down_U']='U2' #user site down\n",
    "part2['Primer_up']=''\n",
    "part2['Primer_up_Tm']=''\n",
    "part2['Primer_down']=''\n",
    "part2['Primer_down_Tm']=''\n",
    "\n",
    "part3={}\n",
    "part3['name']='VPS13t'\n",
    "part3['seq']='gcgcgctgcggatatttctaaggcatggtcgtgcggagctacaataatacgattgaattatagctacatagtgtacaaaagcgggtatatactttcatatgtga'\n",
    "part3['need_reverse']=1#if parts needs to be reversed:1 else:0\n",
    "part3['Primer_up_U']='U2' #user site up\n",
    "part3['Primer_down_U']='GV2' #user site down\n",
    "part3['Primer_up']='' #we dont provide Curran's primer because their TM is very high for VPS13t\n",
    "part3['Primer_up_Tm']=''\n",
    "part3['Primer_down']='' #we dont provide Curran's primer because their TM is very high for VPS13t\n",
    "part3['Primer_down_Tm']=''\n",
    "\n",
    "pCCM011=[part1, part2, part3]"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 15,
   "metadata": {},
   "outputs": [],
   "source": [
    "\n",
    "part17={}\n",
    "part17['name']='pCFB1241_Tef1p_KpAroyC.iso'\n",
    "part17['seq']='gcacacaccatagcttcaaaatgtttctactccttttttactcttccagattttctcggactccgcgcatcgccgtaccacttcaaaacacccaagcacagcatactaaatttcccctctttcttcctctagggtgtcgttaattacccgtactaaaggtttggaaaagaaaaaagagaccgcctcgtttctttttcttcgtcgaaaaaggcaataaaaatttttatcacgtttctttttcttgaaaatttttttttttgatttttttctctttcgatgacctcccattgatatttaagttaataaacggtcttcaatttctcaagtttcagtttcatttttcttgttctattacaactttttttacttcttgctcattagaaagaaagcatagcaatctaatctaagttttaattacaaatctgtcatatctgtcataaaacaatgaccgccccaatccaagatttgagagatgctattgctttgttacaacaacacgacaatcaatacttggaaaccgatcatccagttgatccaaatgctgaattggctggtgtttacagacatattggtgctggtggtactgtaaaaagaccaactagaattggtccagccatgatgttcaacaacattaagggttatccacactccagaatcttggttggtatgcatgcttctagacaaagagcagctttgttgttgggttgtgaagcttctcaattggctttggaagttggtaaagctgttaagaaaccagttgctccagttgttgttccagcttcttctgctccatgtcaagaacaaattttcttggctgatgatccagacttcgatttgagaactttgttgccagctcataccaacactccaattgatgctggtccatttttttgtttgggtttggctttagcttctgatcctgttgatgcttctttgaccgatgttaccattcatagattgtgcgttcaaggtagagatgaattgtctatgtttttggctgccggtagacatatcgaagtttttagacaaaaagctgaagctgctggtaagccattgccaattactattaacatgggtttagatccagccatctacattggtgcttgttttgaagctccaactactccatttggttacaacgaattgggtgttgctggtgctttgagacaaagaccagttgaattggttcaaggtgtttctgttccagaaaaggctattgctagagccgaaatagttatcgaaggtgaattattgccaggtgtcagagttagagaagatcaacatacaaattccggtcatgctatgccagaatttccaggttattgtggtggtgctaatccatctttgccagttattaaggttaaggccgttaccatgagaaacaacgctattttacaaactttggtcggtccaggtgaagaacatacaactttggctggtttgccaaccgaagcttctatttggaatgctgttgaagctgcaattccaggtttcttgcaaaatgtttatgctcatacagctggtggtggtaagttcttgggtatattgcaagtcaagaaaagacaaccagctgacgaaggtagacaaggtcaagctgctttattagctttggctacttactccgaattgaagaatatcatcttggtcgatgaagatgttgatatcttcgattccgatgatattttgtgggctatgactactagaatgcaaggtgatgtttccattactaccattccaggtattagaggtcaccaattagatccatctcaaaccccagaatactccccatcaattagaggtaatggtatctcctgtaagaccattttcgattgcactgttccatgggctttgaagtctcattttgaaagagcaccatttgctgacgttgatcctagaccttttgctccagaatatttcgctagattggaaaagaatcaaggttccgctaagtaa'\n",
    "part17['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "part17['Primer_up_U']='GV1' #user site up\n",
    "part17['Primer_down_U']='U2' #user site down\n",
    "part17['Primer_up']=''\n",
    "part17['Primer_up_Tm']=''\n",
    "part17['Primer_down']=''\n",
    "part17['Primer_down_Tm']=''\n",
    "\n",
    "\n",
    "\n",
    "part18={}\n",
    "part18['name']='IDP1t'\n",
    "part18['seq']='tcgaatttacgtagcccaatctaccacttttttttttcattttttaaagtgttatacttagttatgctctaggataatgaactacttttttttttttttttttactgttatcataaatatatataccttattgttgtttgcaaccgtcggttaattccttatcaaggttccccaagttcggatcattaccatc'\n",
    "part18['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "part18['Primer_up_U']='U2' #user site up\n",
    "part18['Primer_down_U']='GV2' #user site down\n",
    "part18['Primer_up']='tcgaatttacgtagcccaatcta'\n",
    "part18['Primer_up_Tm']=''\n",
    "part18['Primer_down']='gatggtaatgatccgaacttgg'\n",
    "part18['Primer_down_Tm']=''\n",
    "User8=[part17, part18]"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 16,
   "metadata": {},
   "outputs": [],
   "source": [
    "part19={}\n",
    "part19['name']='CCW12p'\n",
    "part19['seq']='tattgatatagtgtttaagcgaatgacagaagattaatttcttggtatgttaggaaagaataaaggagaataagaataattagaacaatgtaggatggaaagaaagattatcaagcatgccgactttatatacttgaacggaggcaaaggatgcaaaattttctcacatttctttctgccgttatgttggaagtaagactcccattatcgcaatactgcaacacgaatatgcaaaatttgctgagttatcgcagatagttgttgcaaagatagcggcgtaggtggccgcgaaatggggaattccaaaacaaacggtttttttactcctgagaaatacttgtacgggataatccagggcctaccacccacgcttcgaggattggcttttatttttttttttttggtggcgttttatttctttcccgctttctgggacttgtgcggagttttgagaggggcgcgcggcaaaggattcccaaaacggaaatcagacgccaatagccagcactcaaagcagttctggacccattccgattttcccatttggttcttgcgcgtgctgattccgacacgcgcgtctataaatagcatgaagtatccgcacaccgcagcgttagtgaggtgagggtggcagcaagctaattcccgcatctggaatctgaactgccccttttggactaaccgtgtggttcatgggtgggcgaagtgcgcaacctcgaaggttttcttttgcgtgtcggattttacatccggcggtagcgcatgatgccatggctggctccagatacatcctcagggcaccagcatctataattagattggcgcaacatggctggctgcactgctgtcttcacttctttctttttccggcaatgaatgatgtatgttttgtggcaaaagggtccgcattgtacctgtttacagttgagattatcgtttttggtagcccttcattacggcataacgtattaagtttcttt'\n",
    "part19['need_reverse']=1 #if parts needs to be reversed:1 else:0\n",
    "part19['Primer_up_U']='GV1' #user site up\n",
    "part19['Primer_down_U']='U1' #user site down\n",
    "part19['Primer_up']=''\n",
    "part19['Primer_up_Tm']=''\n",
    "part19['Primer_down']='aaagaaacttaatacgttatgcc'\n",
    "part19['Primer_down_Tm']=''\n",
    "\n",
    "part20={}\n",
    "part20['name']='pCfB1237_KpAroY.D'\n",
    "part20['seq']='acaatgatctgtccaagatgcgccgacgaaaaaattgaagttatggctacttctccagttaagggtgtttggactgtttatcaatgtcaacactgcttgtacacttggagagatactgaaccattgagaagaacctctagagaacattaccctgaagctttcagaatgacccaaaaggatattgatgaagctccacaagttcctcatgttccaccattattgccagaagataagagataa'\n",
    "part20['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "part20['Primer_up_U']='U1' #user site up\n",
    "part20['Primer_down_U']='U2' #user site down\n",
    "part20['Primer_up']=''\n",
    "part20['Primer_up_Tm']=''\n",
    "part20['Primer_down']=''\n",
    "part20['Primer_down_Tm']=''\n",
    "\n",
    "part21={}\n",
    "part21['name']='PRM9t'\n",
    "part21['seq']='acagaagacgggagacactagcacacaactttaccaggcaaggtatttgacgctagcatgtgtccaattcagtgtcatttatgattttttgtagtaggatataaatatatacagcgctccaaatagtgcggttgccccaaaaacaccacggaacctcatctgttctcgtactttgttgtgacaaagtagctcactgccttattatcacattttcattatgcaacgcttcggaaaatacgatgttgaaaat'\n",
    "part21['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "part21['Primer_up_U']='U2' #user site up\n",
    "part21['Primer_down_U']='GV2' #user site down\n",
    "part21['Primer_up']='' #we dont provide Curran's primer because their TM is very high for PRM9t\n",
    "part21['Primer_up_Tm']=''\n",
    "part21['Primer_down']='' #we dont provide Curran's primer because their TM is very high for PRM9t\n",
    "part21['Primer_down_Tm']=''\n",
    "User9=[part19, part20, part21]"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 17,
   "metadata": {},
   "outputs": [],
   "source": [
    "CCM004={}\n",
    "CCM004['name']='pCCM004(pFBA1:PaAroZ:tHis5)'\n",
    "CCM004['seq']='ACTGGTAGAGAGCGACTTTgtatgccccaattgcgaaacccgcgatatccttctcgattctttagtacccgaccaggacaaggaaaaggaggtcgaaacgtttttgaagaaacaagaggaactacacggaagctctaaagatggcaaccagccagaaactaagaaaatgaagttgatggatccaactggcaccgctggcttgaacaacaataccagccttccaacttctgtaaataacggcggtacgccagtgccaccagtaccgttacctttcggtatacctcctttccccatgtttccaatgcccttcatgcctccaacggctactatcacaaatcctcatcaagctgacgcaagccctaagaaatgaataacaatactgacagtactaaataattgcctacttggcttcacatacgttgcatacgtcgatatagataataatgataatgacagcaggattatcgtaatacgtaatagttgaaaatctcaaaaatgtgtgggtcattacgtaaataatgataggaatgggattcttctatttttcctttttccattctagcagccgtcgggaaaacgtggcatcctctctttcgggctcaattggagtcacgctgccgtgagcatcctctctttccatatctaacaactgagcacgtaaccaatggaaaagcatgagcttagcgttgctccaaaaaagtattggatggttaataccatttgtctgttctcttctgactttgactcctcaaaaaaaaaaaatctacaatcaacagatcgcttcaattacgccctcacaaaaacttttttccttcttcttcgcccacgttaaattttatccctcatgttgtctaacggatttctgcacttgatttattataaaaagacaaagacataatacttctctatcaatttcagttattgttcttccttgcgttattcttctgttcttctttttcttttgtcatatataacCATAACCAAGTAATACATATTCAAAAGCCCTCTtACAATGCCATCCAAGTTGGccattacctctatgtctttgggtagatgttatgccggtcattctttcactactaagttggatatggctagaaagtacggttaccaaggtttggaattattccatgaagatttggctgatgtcgcctatagattgtctggtgaaactccatctccatgtggtccatcaccagctgctcaattgtctgctgctagacaaattttgagaatgtgccaagtcagaaacatcgaaatcgtttgcttgcaaccattctctcaatacgatggtttgttggatagagaagaacacgaaagaagattggaacaattggaattctggatcgaattggcccatgaattggataccgatattattcaaattccagccaacttcttgccagccgaagaagttactgaagatatctctttgattgtctccgacttgcaagaagtagctgatatgggtttacaagctaacccaccaattagattcgtttacgaagctttgtgttggtccactagagttgatacttgggaaagatcttgggaagttgttcaaagagttaacagaccaaacttcggtgtttgtttggacacttttaacattgccggtagagtttatgctgatccaactgttgcttctggtagaactccaaatgctgaagaagctatcagaaagtccattgccagattggttgaaagagttgacgtttccaaggttttctacgttcaagttgttgatgccgaaaagttgaagaaaccattggttccaggtcacagattctatgatccagaacaaccagctagaatgtcttggtctagaaactgcagattattctacggtgaaaaggatagaggtgcttacttgccagtaaaagaaattgcttgggcttttttcaacggtttgggttttgaaggttgggtttccttagaattattcaacagaagaatgtccgataccggttttggtgttccagaagaattagctagaagaggtgctgtttcttgggctaaattggttagagatatgaagatcaccgttgactctccaactcaacaacaagctacacaacaacctatcagaatgttgtCTTTGTCAGCTGCTTTGTGAACACAGGCtATAGATTAATTTAAACAGTATATGTACAGTTTtatatatatatatatatatatatacatatataaagaaacctgtgcgttttttgtattttcaaatacatttagttttggcgccacttcTATAAAAGGTCTCATGATATTGTTAC'\n",
    "CCM004['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "CCM004['Primer_up_U']='GV1' #user site up\n",
    "CCM004['Primer_down_U']='U1'#user site down\n",
    "CCM004['Primer_up']=''\n",
    "CCM004['Primer_up_Tm']=''\n",
    "CCM004['Primer_down']=''\n",
    "CCM004['Primer_down_Tm']=''\n",
    "\n",
    "CCM007={}\n",
    "CCM007['name']='pCCM007(pTDH3:KpAroyB:tVPS13)'\n",
    "CCM007['seq']='ATAAAAAACACGCTTTTTCAgttcgagtttatcattatcaatactgccatttcaaagaatacgtaaataattaatagtagtgattttcctaactttatttagtcaaaaaattagccttttaattctgctgtaacccgtacatgcccaaaatagggggcgggttacacagaatatataacatcgtaggtgtctgggtgaacagtttattcctggcatccactaaatataatggagcccgctttttaagctggcatccagaaaaaaaaagaatcccagcaccaaaatattgttttcttcaccaaccatcagttcataggtccattctcttagcgcaactacagagaacaggggcacaaacaggcaaaaaacgggcacaacctcaatggagtgatgcaacctgcctggagtaaatgatgacacaaggcaattgacccacgcatgtatctatctcattttcttacaccttctattaccttctgctctctctgatttggaaaaagctgaaaaaaaaggttgaaaccagttccctgaaattattcccctacttgactaataagtatataaagacggtaggtattgattgtaattctgtaaatctatttcttaaacttcttaaattctacttttatagttagtcttttttttagttttaaaacaccaagaacttagtttcgaataaacacacataaacaaacaaaagtgcaggtagtgcaggtaaaacaatgaagttgatcatcggtatgactggtgctacaggtgctccattgggtgttgctttgttgcaagctttgagagatatgccagaagttgaaacccatttggttatgtctaaatgggctaagaccaccattgaattggaaactccatggactgctagagaagttgctgctttggctgatttttctcattctccagctgatcaagctgctactatttcttctggttctttcagaactgatggtatgatcgttattccatgctctatgaaaaccttggctggtattagagctggttatgctgaaggtttggttggtagagctgctgatgttgttttgaaagaaggtagaaagttggtcttggtcccaagagaaatgccattgtctactatccatttggaaaacatgttggccttgtctagaatgggtgtagctatggttccaccaatgccagcttattacaatcatccagaaaccgttgatgacatcaccaaccatatagttaccagagttttggaccaattcggtttggattatcacaaagctagaagatggaacggtttgagaactgctgaacaattcgctcaagaaattgaatgaatCGCACGCATTCCGTTGACACAGGCtTCACATATGAAAGTATATACCCGcttttgtacactatgtagctataattcaatcgtattattgtagctccgcacgaccatgccttagaaataTCCGCAGCGCGC'\n",
    "CCM007['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "CCM007['Primer_up_U']='U1' #user site up\n",
    "CCM007['Primer_down_U']='U2'#user site down\n",
    "CCM007['Primer_up']=''\n",
    "CCM007['Primer_up_Tm']=''\n",
    "CCM007['Primer_down']=''\n",
    "CCM007['Primer_down_Tm']=''\n",
    "\n",
    "CCM008={}\n",
    "CCM008['name']='pCCM008(pTEF1:Aroy.Ciso:tIDP1)'\n",
    "CCM008['seq']='GCACACACCATAGCTTCAAaatgtttctactccttttttactcttccagattttctcggactccgcgcatcgccgtaccacttcaaaacacccaagcacagcatactaaatttcccctctttcttcctctagggtgtcgttaattacccgtactaaaggtttggaaaagaaaaaagagaccgcctcgtttctttttcttcgtcgaaaaaggcaataaaaatttttatcacgtttctttttcttgaaaatttttttttttgatttttttctctttcgatgacctcccattgatatttaagttaataaacggtcttcaatttctcaagtttcagtttcatttttcttgttctattacaactttttttacttcttgctcattagaaagaaagcatagcaatctaatctaagttttaattacaatctgtcataaaacaatgaccgccccaatccaagatttgagagatgctattgctttgttacaacaacacgacaatcaatacttggaaaccgatcatccagttgatccaaatgctgaattggctggtgtttacagacatattggtgctggtggtactgtaaaaagaccaactagaattggtccagccatgatgttcaacaacattaagggttatccacactccagaatcttggttggtatgcatgcttctagacaaagagcagctttgttgttgggttgtgaagcttctcaattggctttggaagttggtaaagctgttaagaaaccagttgctccagttgttgttccagcttcttctgctccatgtcaagaacaaattttcttggctgatgatccagacttcgatttgagaactttgttgccagctcataccaacactccaattgatgctggtccatttttttgtttgggtttggctttagcttctgatcctgttgatgcttctttgaccgatgttaccattcatagattgtgcgttcaaggtagagatgaattgtctatgtttttggctgccggtagacatatcgaagtttttagacaaaaagctgaagctgctggtaagccattgccaattactattaacatgggtttagatccagccatctacattggtgcttgttttgaagctccaactactccatttggttacaacgaattgggtgttgctggtgctttgagacaaagaccagttgaattggttcaaggtgtttctgttccagaaaaggctattgctagagccgaaatagttatcgaaggtgaattattgccaggtgtcagagttagagaagatcaacatacaaattccggtcatgctatgccagaatttccaggttattgtggtggtgctaatccatctttgccagttattaaggttaaggccgttaccatgagaaacaacgctattttacaaactttggtcggtccaggtgaagaacatacaactttggctggtttgccaaccgaagcttctatttggaatgctgttgaagctgcaattccaggtttcttgcaaaatgtttatgctcatacagctggtggtggtaagttcttgggtatattgcaagtcaagaaaagacaaccagctgacgaaggtagacaaggtcaagctgctttattagctttggctacttactccgaattgaagaatatcatcttggtcgatgaagatgttgatatcttcgattccgatgatattttgtgggctatgactactagaatgcaaggtgatgtttccattactaccattccaggtattagaggtcaccaattagatccatctcaaaccccagaatactccccatcaattagaggtaatggtatctcctgtaagaccattttcgattgcactgttccatgggctttgaagtctcattttgaaagagcaccatttgctgacgttgatcctagaccttttgctccagaatatttcgctagattggaaaagAATCAAGGTTCCGCTAAGTAAACACAGGCtTCGAATTTACGTAGCCCAATCTAccacttttttttttcattttttaaagtgttatacttagttatgctctaggataatgaactacttttttttttttttttttactgttatcataaatatatataccttattgttgtttgcaaccgtcggttaattccttatcaaggttccCCAAGTTCGGATCATTACCATC'\n",
    "CCM008['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "CCM008['Primer_up_U']='U2' #user site up\n",
    "CCM008['Primer_down_U']='GV2'#user site down\n",
    "CCM008['Primer_up']=''\n",
    "CCM008['Primer_up_Tm']=''\n",
    "CCM008['Primer_down']=''\n",
    "CCM008['Primer_down_Tm']=''\n",
    "\n",
    "pCCM012_13_14_15=[CCM004,CCM007,CCM008]"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 18,
   "metadata": {},
   "outputs": [],
   "source": [
    "CCM009={}\n",
    "CCM009['name']='pCCM009(pCCW12:AroYD:tPRM9)'\n",
    "CCM009['seq']='AAAGAAACTTAATACGTTATGCCgtaatgaagggctaccaaaaacgataatctcaactgtaaacaggtacaatgcggacccttttgccacaaaacatacatcattcattgccggaaaaagaaagaagtgaagacagcagtgcagccagccatgttgcgccaatctaattatagatgctggtgccctgaggatgtatctggagccagccatggcatcatgcgctaccgccggatgtaaaatccgacacgcaaaagaaaaccttcgaggttgcgcacttcgcccacccatgaaccacacggttagtccaaaaggggcagttcagattccagatgcgggaattagcttgctgccaccctcacctcactaacgctgcggtgtgcggatacttcatgctatttatagacgcgcgtgtcggaatcagcacgcgcaagaaccaaatgggaaaatcggaatgggtccagaactgctttgagtgctggctattggcgtctgatttccgttttgggaatcctttgccgcgcgcccctctcaaaactccgcacaagtcccagaaagcgggaaagaaataaaacgccaccaaaaaaaaaaaaataaaagccaatcctcgaagcgtgggtggtaggccctggattatcccgtacaagtatttctcaggagtaaaaaaaccgtttgttttggaattccccatttcgcggccacctacgccgctatctttgcaacaactatctgcgataactcagcaaattttgcatattcgtgttgcagtattgcgataatgggagtcttacttccaacataacggcagaaagaaatgtgagaaaattttgcatcctttgcctccgttcaagtatataaagtcggcatgcttgataatctttctttccatcctacattgttctaattattcttattctcctttattctttcctaacataccaagaaattaatcttctgTCATTCGCTTAAACACTATATCAATAAGCCCTCTtACAATGATCTGTCCAAGATGCgccgacgaaaaaattgaagttatggctacttctccagttaagggtgtttggactgtttatcaatgtcaacactgcttgtacacttggagagatactgaaccattgagaagaacctctagagaacattaccctgaagctttcagaatgacccaaaaggatattgatgaagctccacaagttcctcatgttccacCATTATTGCCAGAAGATAAGAGATAAACACAGGCtACAGAAGACGGGAGACACtagcacacaactttaccaggcaaggtatttgacgctagcatgtgtccaattcagtgtcatttatgattttttgtagtaggatataaatatatacagcgctccaaatagtgcggttgccccaaaaacaccacggaacctcatctgttctcgtactttgttgtgacaaagtagctcactgccttattatcacattttcattatgcaacgctTCGGAAAATACGATGTTGAAAAT'\n",
    "CCM009['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "CCM009['Primer_up_U']='GV1' #user site up\n",
    "CCM009['Primer_down_U']='U1'#user site down\n",
    "CCM009['Primer_up']=''\n",
    "CCM009['Primer_up_Tm']=''\n",
    "CCM009['Primer_down']=''\n",
    "CCM009['Primer_down_Tm']=''\n",
    "\n",
    "CCM005={}\n",
    "CCM005['name']='pCCM005(pTPI1:NrsR:LSC2t)'\n",
    "CCM005['seq']='tAAGGATGAGCCAAGAATAAGGgaacaagattttggtaatttccaaaaaatcaatagcatgcaggacgttatgaagaagagatctacgtatggtcatttcttcttcagattccctcatggagaaagtgcggcagatgtatatgacagagtcgccagtttccaagagactttattcaggcacttccatgataggcaagagagaagacccagagatgttgttgtcctagttacacatggtatttattccagagtattcctgatgaaatggtttagatggacatacgaagagtttgaatcgtttaccaatgttcctaacgggagcgtaatggtgatggaactggacgaatccatcaatagatacgtcctgaggaccgtgctacccaaatggactgattgtgagggagacctaactacatagtgtttaaagattacggatatttaacttacttagaataatgccatttttttgagttataataatcctacgttagtgtgagcgggatttaaactgtgaggaccttaatacattcagacacttctgcggtatcaccctacttattcccttcgagattatatctaggaacccatcaggttggtggaagattacccgttctaagacttttcagcttcctctattgatgttacacctggacaccccttttctggcatccagtttttaatcttcagtggcatgtgagattctccgaaattaattaaagcaatcacacaattctctcggataccacctcggttgaaactgacaggtggtttgttacgcatgctaatgcaaaggagcctatatacctttggctcggctgctgtaacagggaatataaagggcagcataatttaggagtttagtgaacttgcaacatttactattttcccttcttacgtaaatatttttctttttaattctaaatcaatctttttcaattttttgtttgtattcttttcttgcttaaatctataACTACAAAAAACACATACATAAACTAAAAAGCCCTCTtACCATGGGTACCACTCTtgacgacacggcttaccggtaccgcaccagtgtcccgggggacgccgaggccatcgaggcactggatgggtccttcaccaccgacaccgttttccgcgtcaccgccaccggggacggcttcaccctgcgggaggtgccggtggacccgcccctgaccaaggtgttccccgacgacgaatcggacgacgaatcggacgacggggaggacggcgacccggactcccggacgttcgtcgcgtacggggacgacggcgacctggcgggcttcgtggtcgtctcgtactccggctggaaccgccggctgaccgtcgaggacatcgaggtcgccccggagcaccgggggcacggggtcgggcgcgcgttgatggggctcgcgacggagttcgcccgcgagcggggcgccgggcacctctggctggaggtcaccaacgtcaacgcaccggcgatccacgcgtaccggcggatggggttcaccctctgcggcctggacaccgccctgtacgacggcaccgcctcggacggcgagcaggcgctctacatgagcaTGCCCTGCCCCTAAACACAGGCtGCTTCTCGAGAAAAACAAAAGAGTTAataataaagtatatatgcttttttactattaataaagtttcttattttccctacatgtatacatacatataaatatactcttttGGAAAAAATTTCTTTTTTTTTTTTTTAATTTT'\n",
    "CCM005['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "CCM005['Primer_up_U']='U1' #user site up\n",
    "CCM005['Primer_down_U']='U2'#user site down\n",
    "CCM005['Primer_up']=''\n",
    "CCM005['Primer_up_Tm']=''\n",
    "CCM005['Primer_down']=''\n",
    "CCM005['Primer_down_Tm']=''\n",
    "\n",
    "CCM006={}\n",
    "CCM006['name']='pCCM006(pCyc1:CaCatA:CPS1t)'\n",
    "CCM006['seq']='tCAGCATTTTCAAAGGTGTgttcttcgtcagacatgttttagtgtgtgaatgaaataggtgtatgttttctttttgctagacaataattaggaacaaggtaagggaactaaagtgtagaataagattaaaaaagaagaacaagttgaaaaggcaagttgaaatttcaagaaaaaagtcaattgaagtacagtaaattgacctgaatatatctgagttccgacaacaatgagtttaccaaagagaacaatggaataggaaactttgaacgaagaaaggaaagcaggaaaggaaaaaatttttaggctcgagaacaatagggcgaaaaaacaggcaacgaacgaacaatggaaaaacgaaaaaaaaaaaaaaaaacacagaaaagaatgcagaaagatgtcaactgaaaaaaaaaaaggtgaacacaggaaaaaaaataaaaaaaaaaaaaaaaaaaggaggacgaaacaaaaaagtgaaaaaaaatgaaaatttttttggaaaaccaagaaatgaattatatttccgtgtgagacgacatcgtcgaatatgattcagggtaacagtattgatgtaatcaatttcctacctgaatctaaaattcccgggagcaagatcaagatgttttcaccgatctttccggtctctttggccggggtttacggacgatggcagaagaccaaagcgccagttcatttggcgagcgttggttggtggatcaagcccacgcgtaggcaatcctcgagcagatccgccaggcgtgtatatatagcgtggatggccaggcaactttagtgctgacacatacaggcatatatatatgtgtgcgacgacacatgatcatatggcatgcatgtgctctgtatgtatataaaactcttgttttcttcttttctctaaatattctttccttatacattaggacctttgcagcataaattactatacttctatagacacacaAACACAAATACACACACTAAATTAATAAGCCCTCTtACAATGTCCCAAGCTTTCAccgaatctgttaagacttctttgggtccaaatgctactccaagagctaaaaagttgattgcctctttggttcaacacgttcatgatttcgctagagaaaaccatttgactaccgaagattggttgtggggtgttgatttcattaacagaattggtcaaatgtccgactccagaagaaacgaaggtattttggtttgcgatatcatcggtttggaaaccttggttgatgctttgactaacgaatccgaacaatctaaccatacctcctctgctattttgggtcctttttacttgccagattctccagtttatccaaacggtggttctatcgttcaaaaggctattccaactgatgttaagtgcttcgttagaggtaaggttactgatactgaaggtaaaccattgggtggtgctcaattggaagtttggcaatgtaattctgctggtttctactctcaacaagctgatcatgatggtccagaattcaatttgagaggtactttcattaccgacgacgaaggtaattactccttcgaatgtttaagaccaacctcctatccaattccatacgatggtcctgctggtgatttgttgaaaatcatggatagacatccaaacagaccatcccatattcattggagagtttctcatccaggttaccatactttgatcacccaaatctatgatgctgaatgtccatacaccaacaacgattctgtttacgctgttaaggatgacatcatcgttcacttcgaaaaggttgataacaaggataaggatttggtcggtaaggtcgaatacaagttggattacgatatttccttggccaccgaatcctctattcaagaagctagagctgctgctaaagctagaCAAGATGCTGAAATCAAGTTGTAAACACAGGCtGCGCAATGATTGAATAGTCAAAgattttttttttttaattttttttttttaattttttttttttttcatagaactttttatttaaataaatcacgtctatatatgtatcagtataacgtaaaaaaaaaaacaccgtcagttaaacaaaacataaataaaaaaaaAAAGAAGTGTCAAATCAAGTGTCAAATC'\n",
    "CCM006['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "CCM006['Primer_up_U']='U2' #user site up\n",
    "CCM006['Primer_down_U']='U3'#user site down\n",
    "CCM006['Primer_up']=''\n",
    "CCM006['Primer_up_Tm']=''\n",
    "CCM006['Primer_down']=''\n",
    "CCM006['Primer_down_Tm']=''\n",
    "\n",
    "CCM003={}\n",
    "CCM003['name']='pCCM003(pTDH2:Tkl1:TDH3t)'\n",
    "CCM003['seq']='ATCTAGATCAGAGGGTGGTaaatgaagtgtaatagtattcatttttcttataaatcatcccttccgtgatttatacaaaagaagaggagaatatgctgaatacttggtatattactctacattatactcttatcttgacgggtattctgagcatcttactcagtttcaagatcttttaatgtccaaaaacatttgagccgatctaaatacttctgtgttttcattaatttataaattgtactcttttaagacatggaaagtaccaacatcggttgaaacagtttttcatttacttatggtttattggtttttccagtgaatgattatttgtcgttaccctttcgtaaaagttcaaacacgtttttaagtattgtttagttgctctttcgacatatatgattatccctgcgcggctaaagttaaggatgcaaaaaacataagacaactgaagttaatttacgtcaattaagttttccagggtaatgatgttttgggcttccactaattcaataagtatgtcatgaaatacgttgtgaagagcatccagaaataatgaaaagaaacaacgaaactgggtcggcctgttgtttcttttctttaccacgtgatctgcggcatttacaggaagtcgcgcgttttgcgcagttgttgcaacgcagctacggctaacaaagcctagtggaactcgactgatgtgttagggcctaaaactggtggtgacagctgaagtgaactattcaatccaatcatgtcatggctgtcacaaagaccttgcggaccgcacgtacgaacacatacgtatgctaatatgtgttttgatagtacccagtgatcgcagacctgcaatttttttgtaggtttggaagaatatataaaggttgcactcattcaagatagtttttttcttgtgtgtctattcattttattattgtttgtttaaatgttaaaaaaaccaagaacttagtttcaaaTTAAATTCATCACACAAACAAACAAAACAAAAGCCCTCTtACAATGACTCAATTCACTGACATTGATaagctagccgtctccaccataagaattttggctgtggacaccgtatccaaggccaactcaggtcacccaggtgctccattgggtatggcaccagctgcacacgttctatggagtcaaatgcgcatgaacccaaccaacccagactggatcaacagagatagatttgtcttgtctaacggtcacgcggtcgctttgttgtattctatgctacatttgactggttacgatctgtctattgaagacttgaaacagttcagacagttgggttccagaacaccaggtcatcctgaatttgagttgccaggtgttgaagttactaccggtccattaggtcaaggtatctccaacgctgttggtatggccatggctcaagctaacctggctgccacttacaacaagccgggctttaccttgtctgacaactacacctatgttttcttgggtgacggttgtttgcaagaaggtatttcttcagaagcttcctccttggctggtcatttgaaattgggtaacttgattgccatctacgatgacaacaagatcactatcgatggtgctaccagtatctcattcgatgaagatgttgctaagagatacgaagcctacggttgggaagttttgtacgtagaaaatggtaacgaagatctagccggtattgccaaggctattgctcaagctaagttatccaaggacaaaccaactttgatcaaaatgaccacaaccattggttacggttccttgcatgccggctctcactctgtgcacggtgccccattgaaagcagatgatgttaaacaactaaagagcaaattcggtttcaacccagacaagtcctttgttgttccacaagaagtttacgaccactaccaaaagacaattttaaagccaggtgtcgaagccaacaacaagtggaacaagttgttcagcgaataccaaaagaaattcccagaattaggtgctgaattggctagaagattgagcggccaactacccgcaaattgggaatctaagttgccaacttacaccgccaaggactctgccgtggccactagaaaattatcagaaactgttcttgaggatgtttacaatcaattgccagagttgattggtggttctgccgatttaacaccttctaacttgaccagatggaaggaagcccttgacttccaacctccttcttccggttcaggtaactactctggtagatacattaggtacggtattagagaacacgctatgggtgccataatgaacggtatttcagctttcggtgccaactacaaaccatacggtggtactttcttgaacttcgtttcttatgctgctggtgccgttagattgtccgctttgtctggccacccagttatttgggttgctacacatgactctatcggtgtcggtgaagatggtccaacacatcaacctattgaaactttagcacacttcagatccctaccaaacattcaagtttggagaccagctgatggtaacgaagtttctgccgcctacaagaactctttagaatccaagcatactccaagtatcattgctttgtccagacaaaacttgccacaattggaaggtagctctattgaaagcgcttctaagggtggttacgtactacaagatgttgctaacccagatattattttagtggctactggttccgaagtgtctttgagtgttgaagctgctaagactttggccgcaaagaacatcaaggctcgtgttgtttctctaccagatttcttcacttttgacaaacaacccctagaatacagactatcagtcttaccagacaacgttccaatcatgtctgttgaagttttggctaccacatgttggggcaaatacgctcatcaatccttcggtattgacagatttggtgcctccggtaaggcaccagaagtcttcaagttcttcggtttcaccccagaaggtgttgctgaaagagctcaaaagaccattgcattctataagggtgacaagctaatttcTCCTTTGAAAAAAGCTTTCTGAACACAGGCtGTGAATTTACTTTAAATCTTGCATTTAAATAAATTTTCTTTTTATAGCTTTATGACTTAGTTTCAATTTATATACTATTTTAATGACATTTTCGATTCATTGATTGAAAGCTTTGTGTTTTTTCTTGATGCGCTATTGCATTGTTCTTGTCTTTTTCGCCACATGTAATATCTGTAGTAGATACCTGATACATTGTGGATGCTGAGTGAAATTTTAGTTAATAATGGAGGCGCTCTTAATAATTTTGGGGATATTGGCTTTTTTTTTTAAAGTTTACAAATGAATTTTTTCCGCCAGGATAACGATTCTGAAGTTACTCTTAGCGTTCCTATCGGTACAGCCATCAAATCATGCCTATAAATCATGCCTATATTTGCGTGCAGTCAGTATCATCTACATGAAAAAAACTCCCGCAATTTCTTATAGAATACGTTGAAAATTAAATGTACGCGCCAAGATAAGATAACATATATCTAGATGCAGTAATATACACAGATTCCCGCGGACGTGGGAAGGAAAAAATTAGATAACAAAATCTGAGTGATATGGAAATTCCGCTGTATAGCTCATATCTTTCCCTACCTGCAGA'\n",
    "CCM003['need_reverse']=0#if parts needs to be reversed:1 else:0\n",
    "CCM003['Primer_up_U']='U3' #user site up\n",
    "CCM003['Primer_down_U']='GV2'#user site down\n",
    "CCM003['Primer_up']=''\n",
    "CCM003['Primer_up_Tm']=''\n",
    "CCM003['Primer_down']=''\n",
    "CCM003['Primer_down_Tm']=''\n",
    "pCCM016=[CCM009,CCM005,CCM006,CCM003]\n"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 15,
   "metadata": {},
   "outputs": [],
   "source": [
    "CCM001={}\n",
    "CCM001['name']='pCCM001(pCYC_benO:yEGFP:Cyc1t)'\n",
    "CCM001['seq']='GATCCAGGCAACTTTAGTGCtgacacataatactccataggtattttattatacaaataatgtgtttgaacttattaaaacattcttttaaggtataaacaacaggcatatatatatgtgtgcgacgacacatgatcatatggcatgcatgtgctctgtatgtatataaaactcttgttttcttcttttctctaaatattctttccttatacattaggacctttgcagcataaattactatacttctatagacacacaaacacaaatacacacactaaattaataatctgtcataaaacaatgtctaaaggtgaagaattattcactggtgttgtcccaattttggttgaattagatggtgatgttaatggtcacaaattttctgtctccggtgaaggtgaaggtgatgctacttacggtaaattgaccttaaaatttatttgtactactggtaaattgccagttccatggccaaccttagtcactactttcggttatggtgttcaatgttttgcgagatacccagatcatatgaaacaacatgactttttcaagtctgccatgccagaaggttatgttcaagaaagaactatttttttcaaagatgacggtaactacaagaccagagctgaagtcaagtttgaaggtgataccttagttaatagaatcgaattaaaaggtattgattttaaagaagatggtaacattttaggtcacaaattggaatacaactataactctcacaatgtttacatcatggctgacaaacaaaagaatggtatcaaagttaacttcaaaattagacacaacattgaagatggttctgttcaattagctgaccattatcaacaaaatactccaattggtgatggtccagtcttgttaccagacaaccattacttatccactcaatctgccttatccaaagatccaaacgaaaagagagaccacatggtcttgttagaatttgttactgctgctggtattacccatggtatggatgaattgtacaaataaatcgcgtgcattcatccgctctaaccgaaaaggaaggagttagacaacctgaagtctaggtccctatttatttttttatagttatgttagtattaagaacgttatttatatttcaaatttttcttttttttctgtacagacgcgtgtacgcatgtaacattatactgaaaaccttgcttgagaaggttttgggacgctcgaAGATCGCGTCAGCTGAA'\n",
    "CCM001['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "CCM001['Primer_up_U']='GV1' #user site up\n",
    "CCM001['Primer_down_U']='U1'#user site down\n",
    "CCM001['Primer_up']=''\n",
    "CCM001['Primer_up_Tm']=''\n",
    "CCM001['Primer_down']=''\n",
    "CCM001['Primer_down_Tm']=''\n",
    "\n",
    "#this part also works for pCCM010\n",
    "CCM002={}\n",
    "CCM002['name']='pCCM002(tADH1::BenM::pRev1)'\n",
    "CCM002['seq']='gcgacctcatgctatacctgagaaagcaacctgacctacaggaaagagttactcaagaataagaattttcgttttaaaacctaagagtcactttaaaatttgtatacacttattttttttataacttatttaataataaaaatcataaatcataagaaattcgcttatttagaagtgtcaacaacgtatctaccaacggaatgCGTGCGAtGAATGCGGCCGCTTtaccaatttggtggttcagtaaaaccttcgtaggcgtagatttgtctgatggtttcgtataatgagtagatgtaggtggattcttccatgtttctaacagcaatgtagattggggtaatagcatctggatctaataatgggacgtaggacaagttgaataattgaatggattgggtagaagctggaaccaatgaaataccttcaccagcagcaaccaaacccaaggccaattggacttctctaacttcgttaatcttggtaggttccaaaccatggtcagagaagatgttcataacatgagtagagaagtttggctttggagaagatgggtacaacaagatcttttcgtcgatcaaatcgttcaagtgaacacccttatccttcatttgattcaatggatgggaagcatgaacagcaaccatcaatctttcgtttctcaacaaggttctcttgatggctggatcagaaatcttcaatctaccaaaaccagcgtcaattctaccttctttcaaagcttcggtttgagccttagtacccatttcgtataattcgattctcaagtttggatgagcttgtctgtacaagtggataattcttggcaacaaaccgaacaacaaggaaccaacaaaaccgattctaatggtcttttcaacagaggcaattctcttggtcatactgaccatttgatcaacgttgcttagcaacttgatggcgtattggtaaaagaaatgaccttctggagtagtcttaactggtctggaacctctttccaataattggatacccaattcttcttccaagttttggatttgtctggataatggtggttgagcaatacacaacttgtcggcagcctttgtaaaagattgttcttcaacaacggcaacgaagtatctcaagtgtctcaattccattgttttatgacagatcgctggatatgcctagaaatgcagtatcgatttattttgagaaaatctgtcaactcgttacttctttgatagggaaatctttagttaaatttgtttaaaattaagaaccaggtctttagactatgccaaaaggtattggtgcttaaaggttgccgtaaactctaactctatatatctctaagccaatgtttaagttgatgcgatgtgcatgatcacttcacatttttttcatgattttgttttcttccttagctacttttttccgtcttggtctgtgtaaaaaaaaaatcgcttcactgaagaaaaatataaataaaaaaaggacatcagcaagttcgtagtcaataaatgtgatgagtaatttgcgataatctaccccaactcaaagcattcaattcaaatgaaagaagagttcgtgaattttaatctggcttacagattggatcatagcttctaatatatcttcctttttattttaaagtgattaaaatatgtggatgtctgcattaaggtgaaggcagactatagtgaaaaatagtagcaaaggcttcatataatgaactacctagttgtattcaattggaatctgtgaacgcggcattaaaagaatatttgagctttttttgattgtattaacctgaagctatcacggtttagactcattaaaagaaaaatatccggttgtatcggataacaacaccgctgtagtagcacagttttttgatcatagtttcattatttttttaaatatgcttgtatacagtaatctagttttggacggattattcatgtcagtaagattttactatctactttaaaattatcgaaacagatatcaactatacatccactcattattggcatagcgaaataaaatttatgaataggaaggatataaaacgaaaattgaaatcgtttagtattatatgaaactattagtcttttcgttttcgtcgtcatacttacattcttcttttataaatattgttgtgcctaagaagagctccagcttttgttccctttagtgagggttaattgcgcgcttggcgtaatcatggtcatagctgtttccTGTGTGAAATTGTTATCCGCT'\n",
    "CCM002['need_reverse']=1 #if parts needs to be reversed:1 else:0\n",
    "CCM002['Primer_up_U']='U1' #user site up\n",
    "CCM002['Primer_down_U']='GV2'#user site down\n",
    "CCM002['Primer_up']=''\n",
    "CCM002['Primer_up_Tm']=''\n",
    "CCM002['Primer_down']=''\n",
    "CCM002['Primer_down_Tm']=''\n",
    "\n",
    "pCCM017_18_19_20=[CCM001,CCM002]"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 16,
   "metadata": {},
   "outputs": [],
   "source": [
    "#for mads: insertion of BTX in YER084W\n",
    "\n",
    "YER084W_up={}\n",
    "YER084W_up['name']='YER084W_CDS'\n",
    "YER084W_up['seq']='ggtctttgcagcgctaggagcgtctaaaggtgattctgcgttcaaatgactgcttgcctgcccggtgattttattaagcctggagctggcacctccattgctaaacttcttctgtctcctttcccttagtaacctgcgcttttccgcctctgttaattcagacatatttcctagagctacgttgctgatgctacaaaca'\n",
    "YER084W_up['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "YER084W_up['Primer_up_U']='GV1' #user site up\n",
    "YER084W_up['Primer_down_U']='U1'#user site down\n",
    "YER084W_up['Primer_up']=''\n",
    "YER084W_up['Primer_up_Tm']=''\n",
    "YER084W_up['Primer_down']=''\n",
    "YER084W_up['Primer_down_Tm']=''\n",
    "\n",
    "BTX={}\n",
    "BTX['name']='ANTE113_BTX'\n",
    "BTX['seq']='ATTAAGTCCTCAGCGAGCTCGCATGGAATGCGTGCGATGAGCGACCTCATGCTATACCTGAGAAAGCAACCTGACCTACAGGAAAGAGTTACTCAAGAATAAGAATTTTCGTTTTAAAACCTAAGAGTCACTTTAAAATTTGTATACACTTATTTTTTTTATAACTTATTTAATAATAAAAATCATAAATCATAAGAAATTCGCTTATTTAGAAGTGTCAACAACGTATCTACCAACGGAATGCGTGCGATTTACTCTTCCAACCTTCTTAGCAAGTATTCCACCTCAACTTCCTTAGTGATAGGTATAACAATTTCCTTATAAATTTTTACCAAATATTCTGGAGTAATCCTTCTTTCACCTGACTCGTTGGTAGGGTCCACACCATAGACTTCAGCCTTCTTAGTTAATCTTTCTAGAATCTTTTCTTCAACGGCAGAATTGGTGATATTCTTCATTATCCCCTCGACATCTTTACTTTTGATCAGCTTTGTGTATAGCGGGATATCCGATTGGAACTTGGCTTCAGCAACAAACTTGCCAAAGTGGATTCTCCTACTCAAGCTTTGCAAACATTCTATATCTCTAGTGGCAACAGAACTGAAGTTATTCTTATCATCACCATCTCTTTTCGAAATTAATGGTATAATCTTTTCAATATAAACTTTTTTTATTTTATCATTGTAATTAACTTCTGGGGCATAAGGCGCCAAAATTTGTGGGTAGTTAATGCTCGGTAAGAATGATTTCTGAATCTTGTCAGGAAAGAAGGGAGTTTCATCAGGTGATTCGAATCTTCTGATGCGAGAATGCGCAATTTCAAGATTTGAAAGAGCCCAATCCAAGAAAGATCCTTTAAAATTCGGAATTTCTAAACCTGGATGGTTTGCCTCATAAACTGAAGGACATGTGGCGAAATGCGACCTCTCAATAAATTTGAAGATGATCGAATCCTCCATTCTAACTAATTCATCTCTAATATTTTGTAGATTTAAAACAGTTTCTGGTTTTGTGAAATCCATagacttcgtCCGCTTGTAATTAAAACTTAGATTAGATTGCTATGCTTTCTTTCTAATGAGCAAGAAGTAAAAAAAGTTGTAATAGAACAAGAAAAATGAAACTGAAACTTGAGAAATTGAAGACCGTTTATTAACTTAAATATCAATGGGAGGTCATCGAAAGAGAAAAAAATCAAAAAAAAAAATTTTCAAGAAAAAGAAACGTGATAAAAATTTTTATTGCCTTTTTCGACGAAGAAAAAGAAACGAGGCGGTCTCTTTTTTCTTTTCCAAACCTTTAGTACGGGTAATTAACGACACCCTAGAGGAAGAAAGAGGGGAAATTTAGTATGCTGTGCTTGGGTGTTTTGAAGTGGTACGGCGATGCGCGGAGTCCGAGAAAATCTGGAAGAGTAAAAAAGGAGTAGAAACATTTTGAAGCTATGGTGTGTGCGGCCGGCCTGGAAGTACCTTCAAAGAATGGGGTCTTATCTTGTTTTGCAAGTACCACTGAGCAGGATAATAATAGAAATGATAATATACTATAGTAGAGATAACGTCGATGACTTCCCATACTGTAATTGCTTTTAGTTGTGTATTTTTAGTGTGCAAGTTTCTGTAAATCGATTAATTTTTTTTTCTTTCCTCTTTTTATTAACCTTAATTTTTATTTTAGATTCCTGACTTCAACTCAAGACGCACAGATATTATAACATCTGCATAATAGGCATTTGCAAGAATTACTCGTGAGTAAGGAAAGAGTGAGGAACTATCGCATACCTGCATTTAAAGATGCCGATTTGGGCGCGAATCCTTTATTTTGGCTTCACCCTCATACTATTATCAGGGCCAGAAAAAGGAAGTGTTTCCCTCCTTCTTGAATTGATGTTACCCTCATAAAGCACGTGGCCTCTTATCGAGAAAGAAATTACCGTCGCTCGTGATTTGTTTGCAAAAAGAACAAAACTGAAAAAACCCAGACACGCTCGACTTCCTGTCTTCCTATTGATTGCAGCTTCCAATTTCGTCACACAACAAGGTCCTAGCGACGGCTCACAGGTTTTGTAACAAGCAATCGAAGGTTCTGGAATGGCGGGAAAGGGTTTAGTACCACATGCTATGATGCCCACTGTGATCTCCAGAGCAAAGTTCGTTCGATCGTACTGTTACTCTCTCTCTTTCAAACAGAATTGTCCGAATCGTGTGACAACAACAGCCTGTTCTCACACACTCTTTTCTTCTAACCAAGGGGGTGGTTTAGTTTAGTAGAACCTCGTGAAACTTACATTTACATATATATAAACTTGCATAAATTGGTCAATGCAAGAAATACATATTTGGTCTTTTCTAATTCGTAGTTTTTCAAGTTCTTAGATGCTTTCTTTTTCTCTTTTTTACAGATCATCAAGGAAGTAATTATCTACTTTTTACAACAAATATAAAACAAGGATCCaagtctactATGAGTGAATCTCCAATGTTCGCTGCCAACGGCATGCCAAAGGTAAATCAAGGTGCTGAAGAAGATGTCAGAATTTTAGGTTACGACCCATTAGCTTCTCCAGCTCTCCTTCAAGTGCAAATCCCAGCCACACCAACTTCTTTGGAAACTGCCAAGAGAGGTAGAAGAGAAGCTATAGATATTATTACCGGTAAAGACGACAGAGTTCTTGTCATTGTCGGTCCTTGTTCCATCCATGATCTAGAAGCCGCTCAAGAATACGCTTTGAGATTAAAGAAATTGTCAGATGAATTAAAAGGTGATTTATCCATCATTATGAGAGCATACTTGGAGAAGCCAAGAACAACCGTCGGCTGGAAAGGTCTAATTAATGACCCTGATGTTAACAACACTTTCAACATCAACAAGGGTTTGCAATCCGCTAGACAATTGTTTGTCAACTTGACAAATATCGGTTTGCCAATTGGTTCTGAAATGCTTGATACCATTTCTCCTCAATACTTGGCTGATTTGGTCTCCTTCGGTGCCATTGGTGCCAGAACCACCGAATCTCAACTGCACAGAGAATTGGCCTCCGGTTTGTCTTTCCCAGTTGGTTTCAAGAACGGTACCGATGGTACCTTAAATGTTGCTGTGGATGCTTGTCAAGCCGCTGCTCATTCTCACCATTTCATGGGTGTTACTTTACATGGTGTTGCTGCTATCACCACTACTAAGGGTAACGAACACTGCTTCGTTATTCTAAGAGGTGGTAAAAAGGGTACCAACTACGACGCTAAGTCCGTTGCAGAAGCTAAGGCTCAATTGCCTGCCGGTTCCAACGGTCTAATGATTGACTACTCTCACGGTAACTCCAATAAGGATTTCAGAAACCAACCAAAGGTCAATGACGTTGTTTGTGAGCAAATCGCTAACGGTGAAAACGCCATTACCGGTGTCATGATTGAATCAAACATCAACGAAGGTAACCAAGGCATCCCAGCCGAAGGTAAAGCCGGCTTGAAATATGGTGTTTCCATCACTGATGCTTGTATAGGTTGGGAAACTACTGAAGACGTCTTGAGGAAATTGGCTGCTGCTGTCAGACAAAGAAGAGAAGTTAACAAGAAATAGATTCTGCAGGTAGGGAAAGATATGAGCTATACAGCGGAATTTCCATATCACTCAGATTTTGTTATCTAATTTTTTCCTTCCCACGTCCGCGGGAATCTGTGTATATTACTGCATCTAGATATATGTTATCTTATCTTGGCGCGTACATTTAATTTTCAACGTATTCTATAAGAAATTGCGGGAGTTTTTTTCATGTAGATGATACTGACTGCACGCAAATATAGGCATGATTTATAGGCATGATTTGATGGCTGTACCGATAGGAACGCTAAGAGTAACTTCAGAATCGTTATCCTGGCGGAAAAAATTCATTTGTAAACTTTAAAAAAAAAAGCCAATATCCCCAAAATTATTAAGAGCGCCTCCATTATTAACTAAAATTTCACTCAGCATCCACAATGTATCAGGTATCTACTACAGATATTACATGTGGCGAAAAAGACAAGAACAATGCAATAGCGCATCAAGAAAAAACACAAAGCTTTCAATCAATGAATCGAAAATGTCATTAAAATAGTATATAAATTGAAACTAAGTCATAAAGCTATAAAAAGAAAATTTATTTAAATGCAAGATTTAAAGTAAATTCACTTAGGATCCGTATCTTGGAATTGGGATCAATTTTAAAGGTTTAGTTTTCTGCAAGGCTATACCAAACTTTTCATCCATGTCCAAATCTTTAGGTGAAATGTCTCCCTCTAATTTCCAGTTGAAGAATTGCAATAATGTGGCCAACATCAATGTCAACATCCTTATGGCCAAATTCATACCAGGGCAAATTCTTCTACCAGCACCAAAAGGCAATAAACCAAAATCTCTTCCCTTTACATCAATCTCACAACCTATAAATCTTTCAGGTGAGAAAATATCGGCATTTTGCCAAGCATTTGGATCTCTACCTATGGCCCACAAGTTTACCAAAATTTGAGCATCCTTTGGAACAATGTATCCGTACAATTCTACGTCTGTGTCGGCTTTTCTAGGCAACAAGAAAACTGTAGGTGGGTGCAACCTCAAAGTTTCCTTAATGATTGCTTGCAAGTATGGCAAATTTATTATGTCTGACTCTTGGATTTGCTTATCCTTACCCAATACCTGTTTTATTTCCTCTTGGGCCTTTTCCATCATTTCAGGGTTTCTTATTAACTCAGTCATAACCCACTCAAAGGTAGATGATGTAGTATCAGTACCGGCATCGAAAATATCGACTAACAAGTGGTTAATCTCTCCCATAGTTAATTCATTCTGCTTAAACAATTGCAATAATACGTCCAAGACATCATCTGTAGTTGTAGTAGTTGTAGTTGATGAATCTGGAGCTAACCTCTCACATATTATTCCTTGAAATACTGCAATTAACTTATCAAAAGAACATGCCAACCTTCTTCTAATTCCTGATGGATCGACGCAACCTAAAATTGGGAAGTAATCGGCGTAGTTTGGTTTACCGATGTCTTCCATAATATTCCAAATTAATTCCTTAAACTCTTGAGAGGTATGTGATTTATGATGGGCCAATTCAACAGAAAAGAATAATTTAGACAATAAATTCAAAGAAGTTGTGAAGGCAGCTTTACCAATGTCAACTGCCTGACCCTTCTGAGCGCACTCTTGAACGTATTCATATAATTGCTGAACCTTTGCATGTCTAAAGGTCTGGCATGCATCTAATCTTTGTGGAGACAATAAATGAACAGCAGTTATCTTTCTGAAGTTTCTCCATTTAGGTGAAACAGGCAACCAAGACATAGTCAATTTGTGGTGGTCACCAGCAGTTACTGAATTTGGAATTGTCCTGTTTGATAAAGGGTGATCCTTCTTCAAGAACATCTCTTTGGCGACATCAGCAGATGATACAACAATGGTGGTAACTGATCCTAACCTCAAAGAGATTAAAGGACCGTGGATCTTAGCCAAATTTGCAAATGATCTATGAGGCTTCTTTCCTACTTCCAAGATATTACCAATAATTGGTAAAGGTTTAGGACCTGGAGGTAACAATTTTGTTGTTTGTTGTGAGAATAATAATTTAATAAAATGAAATGAAATAAACCAAATTGCTAATATCATAGCTAATGTTGCATGATCCATTTTTAGTTTATGTATGTGTTTTTTGTAGTTATAGATTTAAGCAAGAAAAGAATACAAACAAAAAATTGAAAAAGATTGATTTAGAATTAAAAAGAAAAATATTTACGTAAGAAGGGAAAATAGTAAATGTTGCAAGTTCACTAAACTCCTAAATTATGCTGCCCTTTATATTCCCTGTTACAGCAGCCGAGCCAAAGGTATATAGGCTCCTTTGCATTAGCATGCGTAACAAACCACCTGTCAGTTTCAACCGAGGTGGTATCCGAGAGAATTGTGTGATTGCTTTAATTAATTTCGGAGAATCTCACATGCCACTGAAGATTAAAAACTGGATGCCAGAAAAGGGGTGTCCAGGTGTAACATCAATAGAGGAAGCTGAAAAGTCTTAGAACGGGTAATCTTCCACCAACCTGATGGGTTCCTAGATATAATCTCGAAGGGAATAAGTAGGGTGATACCGCAGAAGTGTCTGAATGTATTAAGGTCCTCACAGTTTAAATCCCGCTCACACTAACGTAGGATTATTATAACTCAAAAAAATGGCATTATTCTAAGTAAGTTAAATATCCGTAATCTTTAAACAagacttcgTCACCCATGAACCACACGGTTAGTCCAAAAGGGGCAGTTCAGATTCCAGATGCGGGAATTAGCTTGCTGCCACCCTCACCTCACTAACGCTGCGGTGTGCGGATACTTCATGCTATTTATAGACGCGCGTGTCGGAATCAGCACGCGCAAGAACCAAATGGGAAAATCGGAATGGGTCCAGAACTGCTTTGAGTGCTGGCTATTGGCGTCTGATTTCCGTTTTGGGAATCCTTTGCCGCGCGCCCCTCTCAAAACTCCGCACAAGTCCCAGAAAGCGGGAAAGAAATAAAACGCCACCAAAAAAAAAAAAATAAAAGCCAATCCTCGAAGCGTGGGTGGTAGGCCCTGGATTATCCCGTACAAGTATTTCTCAGGAGTAAAAAAACCGTTTGTTTTGGAATTTCCCATTTCGCGGCCACCTACGCCGCTATCTTTGCAACAACTATCTGCGATAACTCAGCAAATTTTGCATATTCGTGTTGCAGTATTGCGATAATGGGAGTCTTACTTCCAACATAACGGCAGAAAGAAATGTGAGAAAATTTTGCATCCTTTGCCTCCGTTCAAGTATATAAAGTCGGCATGCTTGATAATCTTTCTTTCCATCCTACATTGTTCTAATTATTCTTATTCTCCTTTATTCTTTCCTAACATACCAAGAAATTAATCTTCTGTCATTCGCTTAAACACTATATCAATAAAGATCTAAAAATGAAGGGAACCTACTACATCAACCACGGTGACCCATTGATGTACTTGAAGAAGCACATCAAGTTGAGACAATTCTTGGAAGGTTGGCAAGAAAACGTTGTTATCGAAAAGCCAAAGTCTATCTTGATCATCAGTGCACACTGGGACACCAACGTTCCAACCGTTAACTTCGTTGAACACTGTGACACCATCCACGACTTCGACGACTACCCAGACCCATTGTACCAAATCCAATACCGCGCTCCAGGTGCTCCAAACTTGGCTAAGAAGGTTGAAGAATTGCTTAAGGAATCTGGTATGGAATGTGAAATCGACACCAAGAGAGGTTTGGACCACGCTGCTTGGTTCCCATTGATGTTCATGTACCCAGAAGCTAACATCCCAATCTGTGAATTGTCTGTTCAACCATCTAAGGACGGTATCCACCACTACAACGTTGGTAAAGCTTTGTCTCCATTGTTGCAACAAGGTGTTTTGATCATCGGTTCTGGTGGAACCGTTCACCCATCTGACGACACCCCACACTGTCCAAACGGTGTTGCTCCATGGGCTATCGAGTTCGACAACTGGTTGGAAGACGCTTTGCTCTCGGGTAGATACGAAGACGTTAACAACTTCAAGAAGTTGGCTCCAAACTGGGAAATCTCTCACCCAGGTCAAGAACACTTGTACCCATTGCACGTTGCTTTGGGTGCTGCTGGTAAGAACCCAAAGACCCAATTGATCCACCGGTCTTGGGCTGCTAACGGTGTTTTCGGTTACTCTACCTACAACTTCACCCCAACCACCCAAAAGACCGACGGATCCTAAATCGCGTGCATTCATCCGCTCTAACCGAAAAGGAAGGAGTTAGACAACCTGAAGTCTAGGTCCCTATTTATTTTTTTATAGTTATGTTAGTATTAAGAACGTTATTTATATTTCAAATTTTTCTTTTTTTTCTGTACAGACGCGTGTACGCATGTAACATTATACTGAAAACCTTGCTTGAGAAGGTTTTGGGACGCTCGAAGATCGCGTCAGCTGAAGCTTCGTACGCTGCAGGTCGACAATTCTGCAGGTCGACAACCCTTAATATAACTTCGTATAATGTATGCTATACGAAGTTATTAGGTCTAGAGATCTGTTTAGCTTGCCTCGTCCCCGCCGGGTCACCCGGCCAGCGACATGGAGGCCCAGAATACCCTCCTTGACAGTCTTGACGTGCGCAGCTCAGGGGCATGATGTGACTGTCGCCCGTACATTTAGCCCATACATCCCCATGTATAATCATTTGCATCCATACATTTTGATGGCCGCACGGCGCGAAGCAAAAATTACGGCTCCTCGCTGCAGACCTGCGAGCAGGGAAACGCTCCCCTCACAGACGCGTTGAATTGTCCCCACGCCGCGCCCCTGTAGAGAAATATAAAAGGTTAGGATTTGCCACTGAGGTTCTTCTTTCATATACTTCCTTTTAAAATCTTGCTAGGATACAGTTCTCACATCACATCCGAACATAAACAACCATGGGTACCACTCTTGACGACACGGCTTACCGGTACCGCACCAGTGTCCCGGGGGACGCCGAGGCCATCGAGGCACTGGATGGGTCCTTCACCACCGACACCGTCTTCCGCGTCACCGCCACCGGGGACGGCTTCACCCTGCGGGAGGTGCCGGTGGACCCGCCCCTGACCAAGGTGTTCCCCGACGACGAATCGGACGACGAATCGGACGACGGGGAGGACGGCGACCCGGACTCCCGGACGTTCGTCGCGTACGGGGACGACGGCGACCTGGCGGGCTTCGTGGTCGTCTCGTACTCCGGCTGGAACCGCCGGCTGACCGTCGAGGACATCGAGGTCGCCCCGGAGCACCGGGGGCACGGGGTCGGGCGCGCGTTGATGGGGCTCGCGACGGAGTTCGCCCGCGAGCGGGGCGCCGGGCACCTCTGGCTGGAGGTCACCAACGTCAACGCACCGGCGATCCACGCGTACCGGCGGATGGGGTTCACCCTCTGCGGCCTGGACACCGCCCTGTACGACGGCACCGCCTCGGACGGCGAGCAGGCGCTCTACATGAGCATGCCCTGCCCCTAATCAGTACTGACAATAAAAAGATTCTTGTTTTCAAGAACTTGTCATTTGTATAGTTTTTTTATATTGTAGTTGTTCTATTTTAATCAAATGTTAGCGTGATTTATATTTTTTTTCGCCTCGACATCATCTGCCCAGATGCGAAGTTAAGTGCGCAGAAAGTAATATCATGCGTCAATCGTATGTGAATGCTGGTCGCTATACTGCTGTCGATTCGATACTAACGCCGCCATCCAGTGTCGAAAACGAGCTCTCGAGAACCCTTAATATAACTTCGTATAATGTATGCTATACGAAGTTATTAGGTGATATCAGATCCACTAGTG'\n",
    "BTX['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "BTX['Primer_up_U']='U1' #user site up\n",
    "BTX['Primer_down_U']='U2'#user site down\n",
    "BTX['Primer_up']=''\n",
    "BTX['Primer_up_Tm']=''\n",
    "BTX['Primer_down']=''\n",
    "BTX['Primer_down_Tm']=''\n",
    "\n",
    "YER084W_dw={}\n",
    "YER084W_dw['name']='YER084W_CDS'\n",
    "YER084W_dw['seq']='taactcagattttcaagaaacagaatagcacattctctaacatcaagttatcataccaatatataacacaaattatgcataaatagataagtaatatgtccctctgcccctgcctttccctttccagatgctggcatcgaaagtaaggcatgtatttgtatgtaagttttttaattaatagtcttcttctttcttatcag'\n",
    "YER084W_dw['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "YER084W_dw['Primer_up_U']='U2' #user site up\n",
    "YER084W_dw['Primer_down_U']='GV2'#user site down\n",
    "YER084W_dw['Primer_up']=''\n",
    "YER084W_dw['Primer_up_Tm']=''\n",
    "YER084W_dw['Primer_down']=''\n",
    "YER084W_dw['Primer_down_Tm']=''\n",
    "\n",
    "YER084W_KO_BTX=[YER084W_up,BTX,YER084W_dw]"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 9,
   "metadata": {},
   "outputs": [],
   "source": [
    "DOPA1={}\n",
    "DOPA1['name']='ANTE113:ARO7G141S_ARO4K229L_vCYP76AD1'\n",
    "DOPA1['seq']='GCGACCTCATGCTATACCTGAGAAAGCAACCTGACCTACAGGAAAGAGTTACTCAAGAATAAGAATTTTCGTTTTAAAACCTAAGAGTCACTTTAAAATTTGTATACACTTATTTTTTTTATAACTTATTTAATAATAAAAATCATAAATCATAAGAAATTCGCTTATTTAGAAGTGTCAACAACGTATCTACCAACGGAATGCGTGCGATTTACTCTTCCAACCTTCTTAGCAAGTATTCCACCTCAACTTCCTTAGTGATAGGTATAACAATTTCCTTATAAATTTTTACCAAATATTCTGGAGTAATCCTTCTTTCACCTGACTCGTTGGTAGGGTCCACACCATAGACTTCAGCCTTCTTAGTTAATCTTTCTAGAATCTTTTCTTCAACGGCAGAATTGGTGATATTCTTCATTATCCCCTCGACATCTTTACTTTTGATCAGCTTTGTGTATAGCGGGATATCCGATTGGAACTTGGCTTCAGCAACAAACTTGCCAAAGTGGATTCTCCTACTCAAGCTTTGCAAACATTCTATATCTCTAGTGGCAACAGAACTGAAGTTATTCTTATCATCACCATCTCTTTTCGAAATTAATGGTATAATCTTTTCAATATAAACTTTTTTTATTTTATCATTGTAATTAACTTCTGGGGCATAAGGCGCCAAAATTTGTGGGTAGTTAATGCTCGGTAAGAATGATTTCTGAATCTTGTCAGGAAAGAAGGGAGTTTCATCAGGTGATTCGAATCTTCTGATGCGAGAATGCGCAATTTCAAGATTTGAAAGAGCCCAATCCAAGAAAGATCCTTTAAAATTCGGAATTTCTAAACCTGGATGGTTTGCCTCATAAACTGAAGGACATGTGGCGAAATGCGACCTCTCAATAAATTTGAAGATGATCGAATCCTCCATTCTAACTAATTCATCTCTAATATTTTGTAGATTTAAAACAGTTTCTGGTTTTGTGAAATCCATagacttcgtCCGCTTGTAATTAAAACTTAGATTAGATTGCTATGCTTTCTTTCTAATGAGCAAGAAGTAAAAAAAGTTGTAATAGAACAAGAAAAATGAAACTGAAACTTGAGAAATTGAAGACCGTTTATTAACTTAAATATCAATGGGAGGTCATCGAAAGAGAAAAAAATCAAAAAAAAAAATTTTCAAGAAAAAGAAACGTGATAAAAATTTTTATTGCCTTTTTCGACGAAGAAAAAGAAACGAGGCGGTCTCTTTTTTCTTTTCCAAACCTTTAGTACGGGTAATTAACGACACCCTAGAGGAAGAAAGAGGGGAAATTTAGTATGCTGTGCTTGGGTGTTTTGAAGTGGTACGGCGATGCGCGGAGTCCGAGAAAATCTGGAAGAGTAAAAAAGGAGTAGAAACATTTTGAAGCTATGGTGTGTGCGGCCGGCCTGGAAGTACCTTCAAAGAATGGGGTCTTATCTTGTTTTGCAAGTACCACTGAGCAGGATAATAATAGAAATGATAATATACTATAGTAGAGATAACGTCGATGACTTCCCATACTGTAATTGCTTTTAGTTGTGTATTTTTAGTGTGCAAGTTTCTGTAAATCGATTAATTTTTTTTTCTTTCCTCTTTTTATTAACCTTAATTTTTATTTTAGATTCCTGACTTCAACTCAAGACGCACAGATATTATAACATCTGCATAATAGGCATTTGCAAGAATTACTCGTGAGTAAGGAAAGAGTGAGGAACTATCGCATACCTGCATTTAAAGATGCCGATTTGGGCGCGAATCCTTTATTTTGGCTTCACCCTCATACTATTATCAGGGCCAGAAAAAGGAAGTGTTTCCCTCCTTCTTGAATTGATGTTACCCTCATAAAGCACGTGGCCTCTTATCGAGAAAGAAATTACCGTCGCTCGTGATTTGTTTGCAAAAAGAACAAAACTGAAAAAACCCAGACACGCTCGACTTCCTGTCTTCCTATTGATTGCAGCTTCCAATTTCGTCACACAACAAGGTCCTAGCGACGGCTCACAGGTTTTGTAACAAGCAATCGAAGGTTCTGGAATGGCGGGAAAGGGTTTAGTACCACATGCTATGATGCCCACTGTGATCTCCAGAGCAAAGTTCGTTCGATCGTACTGTTACTCTCTCTCTTTCAAACAGAATTGTCCGAATCGTGTGACAACAACAGCCTGTTCTCACACACTCTTTTCTTCTAACCAAGGGGGTGGTTTAGTTTAGTAGAACCTCGTGAAACTTACATTTACATATATATAAACTTGCATAAATTGGTCAATGCAAGAAATACATATTTGGTCTTTTCTAATTCGTAGTTTTTCAAGTTCTTAGATGCTTTCTTTTTCTCTTTTTTACAGATCATCAAGGAAGTAATTATCTACTTTTTACAACAAATATAAAACAAGGATCCaagtctactATGAGTGAATCTCCAATGTTCGCTGCCAACGGCATGCCAAAGGTAAATCAAGGTGCTGAAGAAGATGTCAGAATTTTAGGTTACGACCCATTAGCTTCTCCAGCTCTCCTTCAAGTGCAAATCCCAGCCACACCAACTTCTTTGGAAACTGCCAAGAGAGGTAGAAGAGAAGCTATAGATATTATTACCGGTAAAGACGACAGAGTTCTTGTCATTGTCGGTCCTTGTTCCATCCATGATCTAGAAGCCGCTCAAGAATACGCTTTGAGATTAAAGAAATTGTCAGATGAATTAAAAGGTGATTTATCCATCATTATGAGAGCATACTTGGAGAAGCCAAGAACAACCGTCGGCTGGAAAGGTCTAATTAATGACCCTGATGTTAACAACACTTTCAACATCAACAAGGGTTTGCAATCCGCTAGACAATTGTTTGTCAACTTGACAAATATCGGTTTGCCAATTGGTTCTGAAATGCTTGATACCATTTCTCCTCAATACTTGGCTGATTTGGTCTCCTTCGGTGCCATTGGTGCCAGAACCACCGAATCTCAACTGCACAGAGAATTGGCCTCCGGTTTGTCTTTCCCAGTTGGTTTCAAGAACGGTACCGATGGTACCTTAAATGTTGCTGTGGATGCTTGTCAAGCCGCTGCTCATTCTCACCATTTCATGGGTGTTACTTTACATGGTGTTGCTGCTATCACCACTACTAAGGGTAACGAACACTGCTTCGTTATTCTAAGAGGTGGTAAAAAGGGTACCAACTACGACGCTAAGTCCGTTGCAGAAGCTAAGGCTCAATTGCCTGCCGGTTCCAACGGTCTAATGATTGACTACTCTCACGGTAACTCCAATAAGGATTTCAGAAACCAACCAAAGGTCAATGACGTTGTTTGTGAGCAAATCGCTAACGGTGAAAACGCCATTACCGGTGTCATGATTGAATCAAACATCAACGAAGGTAACCAAGGCATCCCAGCCGAAGGTAAAGCCGGCTTGAAATATGGTGTTTCCATCACTGATGCTTGTATAGGTTGGGAAACTACTGAAGACGTCTTGAGGAAATTGGCTGCTGCTGTCAGACAAAGAAGAGAAGTTAACAAGAAATAGATTCTGCAGGTAGGGAAAGATATGAGCTATACAGCGGAATTTCCATATCACTCAGATTTTGTTATCTAATTTTTTCCTTCCCACGTCCGCGGGAATCTGTGTATATTACTGCATCTAGATATATGTTATCTTATCTTGGCGCGTACATTTAATTTTCAACGTATTCTATAAGAAATTGCGGGAGTTTTTTTCATGTAGATGATACTGACTGCACGCAAATATAGGCATGATTTATAGGCATGATTTGATGGCTGTACCGATAGGAACGCTAAGAGTAACTTCAGAATCGTTATCCTGGCGGAAAAAATTCATTTGTAAACTTTAAAAAAAAAAGCCAATATCCCCAAAATTATTAAGAGCGCCTCCATTATTAACTAAAATTTCACTCAGCATCCACAATGTATCAGGTATCTACTACAGATATTACATGTGGCGAAAAAGACAAGAACAATGCAATAGCGCATCAAGAAAAAACACAAAGCTTTCAATCAATGAATCGAAAATGTCATTAAAATAGTATATAAATTGAAACTAAGTCATAAAGCTATAAAAAGAAAATTTATTTAAATGCAAGATTTAAAGTAAATTCACTTAGGATCCGTATCTTGGAATTGGGATCAATTTTAAAGGTTTAGTTTTCTGCAAGGCTATACCAAACTTTTCATCCATGTCCAAATCTTTAGGTGAAATGTCTCCCTCTAATTTCCAGTTGAAGAATTGCAATAATGTGGCCAACATCAATGTCAACATCCTTATGGCCAAATTCATACCAGGGCAAATTCTTCTACCAGCACCAAAAGGCAATAAACCAAAATCTCTTCCCTTTACATCAATCTCACAACCTATAAATCTTTCAGGTGAGAAAATATCGGCATTTTGCCAAGCATTTGGATCTCTACCTATGGCCCACAAGTTTACCAAAATTTGAGCATCCTTTGGAACAATGTATCCGTACAATTCTACGTCTGTGTCGGCTTTTCTAGGCAACAAGAAAACTGTAGGTGGGTGCAACCTCAAAGTTTCCTTAATGATTGCTTGCAAGTATGGCAAATTTATTATGTCTGACTCTTGGATTTGCTTATCCTTACCCAATACCTGTTTTATTTCCTCTTGGGCCTTTTCCATCATTTCAGGGTTTCTTATTAACTCAGTCATAACCCACTCAAAGGTAGATGATGTAGTATCAGTACCGGCATCGAAAATATCGACTAACAAGTGGTTAATCTCTCCCATAGTTAATTCATTCTGCTTAAACAATTGCAATAATACGTCCAAGACATCATCTGTAGTTGTAGTAGTTGTAGTTGATGAATCTGGAGCTAACCTCTCACATATTATTCCTTGAAATACTGCAATTAACTTATCAAAAGAACATGCCAACCTTCTTCTAATTCCTGATGGATCGACGCAACCTAAAATTGGGAAGTAATCGGCGTAGTTTGGTTTACCGATGTCTTCCATAATATTCCAAATTAATTCCTTAAACTCTTGAGAGGTATGTGATTTATGATGGGCCAATTCAACAGAAAAGAATAATTTAGACAATAAATTCAAAGAAGTTGTGAAGGCAGCTTTACCAATGTCAACTGCCTGACCCTTCTGAGCGCACTCTTGAACGTATTCATATAATTGCTGAACCTTTGCATGTCTAAAGGTCTGGCATGCATCTAATCTTTGTGGAGACAATAAATGAACAGCAGTTATCTTTCTGAAGTTTCTCCATTTAGGTGAAACAGGCAACCAAGACATAGTCAATTTGTGGTGGTCACCAGCAGTTACTGAATTTGGAATTGTCCTGTTTGATAAAGGGTGATCCTTCTTCAAGAACATCTCTTTGGCGACATCAGCAGATGATACAACAATGGTGGTAACTGATCCTAACCTCAAAGAGATTAAAGGACCGTGGATCTTAGCCAAATTTGCAAATGATCTATGAGGCTTCTTTCCTACTTCCAAGATATTACCAATAATTGGTAAAGGTTTAGGACCTGGAGGTAACAATTTTGTTGTTTGTTGTGAGAATAATAATTTAATAAAATGAAATGAAATAAACCAAATTGCTAATATCATAGCTAATGTTGCATGATCCATTTTTAGTTTATGTATGTGTTTTTTGTAGTTATAGATTTAAGCAAGAAAAGAATACAAACAAAAAATTGAAAAAGATTGATTTAGAATTAAAAAGAAAAATATTTACGTAAGAAGGGAAAATAGTAAATGTTGCAAGTTCACTAAACTCCTAAATTATGCTGCCCTTTATATTCCCTGTTACAGCAGCCGAGCCAAAGGTATATAGGCTCCTTTGCATTAGCATGCGTAACAAACCACCTGTCAGTTTCAACCGAGGTGGTATCCGAGAGAATTGTGTGATTGCTTTAATTAATTTCGGAGAATCTCACATGCCACTGAAGATTAAAAACTGGATGCCAGAAAAGGGGTGTCCAGGTGTAACATCAATAGAGGAAGCTGAAAAGTCTTAGAACGGGTAATCTTCCACCAACCTGATGGGTTCCTAGATATAATCTCGAAGGGAATAAGTAGGGTGATACCGCAGAAGTGTCTGAATGTATTAAGGTCCTCACAGTTTAAATCCCGCTCACACTAACGTAGGATTATTATAACTCAAAAAAATGGCATTATTCTAAGTAAGTTAAATATCCGTAATCTTTAAACA'\n",
    "DOPA1['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "DOPA1['Primer_up_U']='GV1' #user site up\n",
    "DOPA1['Primer_down_U']='U1'#user site down\n",
    "DOPA1['Primer_up']=''\n",
    "DOPA1['Primer_up_Tm']=''\n",
    "DOPA1['Primer_down']=''\n",
    "DOPA1['Primer_down_Tm']=''\n",
    "\n",
    "DOPA2={}\n",
    "DOPA2['name']='ANTE113:NatMX'\n",
    "DOPA2['seq']='TAGGTCTAGAGATCTGTTTAGCTTGCCTCGTCCCCGCCGGGTCACCCGGCCAGCGACATGGAGGCCCAGAATACCCTCCTTGACAGTCTTGACGTGCGCAGCTCAGGGGCATGATGTGACTGTCGCCCGTACATTTAGCCCATACATCCCCATGTATAATCATTTGCATCCATACATTTTGATGGCCGCACGGCGCGAAGCAAAAATTACGGCTCCTCGCTGCAGACCTGCGAGCAGGGAAACGCTCCCCTCACAGACGCGTTGAATTGTCCCCACGCCGCGCCCCTGTAGAGAAATATAAAAGGTTAGGATTTGCCACTGAGGTTCTTCTTTCATATACTTCCTTTTAAAATCTTGCTAGGATACAGTTCTCACATCACATCCGAACATAAACAACCATGGGTACCACTCTTGACGACACGGCTTACCGGTACCGCACCAGTGTCCCGGGGGACGCCGAGGCCATCGAGGCACTGGATGGGTCCTTCACCACCGACACCGTCTTCCGCGTCACCGCCACCGGGGACGGCTTCACCCTGCGGGAGGTGCCGGTGGACCCGCCCCTGACCAAGGTGTTCCCCGACGACGAATCGGACGACGAATCGGACGACGGGGAGGACGGCGACCCGGACTCCCGGACGTTCGTCGCGTACGGGGACGACGGCGACCTGGCGGGCTTCGTGGTCGTCTCGTACTCCGGCTGGAACCGCCGGCTGACCGTCGAGGACATCGAGGTCGCCCCGGAGCACCGGGGGCACGGGGTCGGGCGCGCGTTGATGGGGCTCGCGACGGAGTTCGCCCGCGAGCGGGGCGCCGGGCACCTCTGGCTGGAGGTCACCAACGTCAACGCACCGGCGATCCACGCGTACCGGCGGATGGGGTTCACCCTCTGCGGCCTGGACACCGCCCTGTACGACGGCACCGCCTCGGACGGCGAGCAGGCGCTCTACATGAGCATGCCCTGCCCCTAATCAGTACTGACAATAAAAAGATTCTTGTTTTCAAGAACTTGTCATTTGTATAGTTTTTTTATATTGTAGTTGTTCTATTTTAATCAAATGTTAGCGTGATTTATATTTTTTTTCGCCTCGACATCATCTGCCCAGATGCGAAGTTAAGTGCGCAGAAAGTAATATCATGCGTCAATCGTATGTGAATGCTGGTCGCTATACTGCTGTCGATTCGATACTAACGCCGCCATCCAGTGTCGAAAACGAGCTCTCGAGAACCCTTAAT'\n",
    "DOPA2['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "DOPA2['Primer_up_U']='U1' #user site up\n",
    "DOPA2['Primer_down_U']='GV2'#user site down\n",
    "DOPA2['Primer_up']=''\n",
    "DOPA2['Primer_up_Tm']=''\n",
    "DOPA2['Primer_down']=''\n",
    "DOPA2['Primer_down_Tm']=''\n",
    "\n",
    "DOPAs=[DOPA1,DOPA2]"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 69,
   "metadata": {},
   "outputs": [],
   "source": [
    "### Design the primers for addind CDSs to the BTX pathway + making the KO:\n",
    "###up regions (promoter): 1000bp upstream of CDS unless another sequeunce is there\n",
    "### upstream is 500bp regardless of presence of other CDS (might include the end of another gene)\n",
    "### for PDX3 promoter is close to snoRNA, so only took 240bp long promoter\n",
    "### for PDX3 terminator is close to start of other CDS so took a shorter terminator\n",
    "##down region (terminator): 500dp dwnstream of CDS unless another sequeunce is there\n",
    "\n",
    "#for the KO primers (reverse of up and forward of down) to avoid nesting promoter>>promoter::KANMX::terminator>>terminator\n",
    "#end up region so that total length is 200bp\n",
    "#start dwn region so that total length is 200bp\n",
    "\n",
    "#YDL119C =HEM25\n",
    "#KGD2 v\n",
    "#PPA1 = VMA16\n",
    "#CAX4\n",
    "#Pro2\n",
    "#GLR1\n",
    "#YSY6\n",
    "#PDX3\n",
    "\n",
    "#BTX pathway will be annealed straight into pCCM021 to be flanked by XII-up and  HomoB\n",
    "BTX={}\n",
    "BTX['name']='ANTE113_BTX'\n",
    "BTX['seq']='GCGACCTCATGCTATACCTGAGAAAGCAACCTGACCTACAGGAAAGAGTTACTCAAGAATAAGAATTTTCGTTTTAAAACCTAAGAGTCACTTTAAAATTTGTATACACTTATTTTTTTTATAACTTATTTAATAATAAAAATCATAAATCATAAGAAATTCGCTTATTTAGAAGTGTCAACAACGTATCTACCAACGGAATGCGTGCGATTTACTCTTCCAACCTTCTTAGCAAGTATTCCACCTCAACTTCCTTAGTGATAGGTATAACAATTTCCTTATAAATTTTTACCAAATATTCTGGAGTAATCCTTCTTTCACCTGACTCGTTGGTAGGGTCCACACCATAGACTTCAGCCTTCTTAGTTAATCTTTCTAGAATCTTTTCTTCAACGGCAGAATTGGTGATATTCTTCATTATCCCCTCGACATCTTTACTTTTGATCAGCTTTGTGTATAGCGGGATATCCGATTGGAACTTGGCTTCAGCAACAAACTTGCCAAAGTGGATTCTCCTACTCAAGCTTTGCAAACATTCTATATCTCTAGTGGCAACAGAACTGAAGTTATTCTTATCATCACCATCTCTTTTCGAAATTAATGGTATAATCTTTTCAATATAAACTTTTTTTATTTTATCATTGTAATTAACTTCTGGGGCATAAGGCGCCAAAATTTGTGGGTAGTTAATGCTCGGTAAGAATGATTTCTGAATCTTGTCAGGAAAGAAGGGAGTTTCATCAGGTGATTCGAATCTTCTGATGCGAGAATGCGCAATTTCAAGATTTGAAAGAGCCCAATCCAAGAAAGATCCTTTAAAATTCGGAATTTCTAAACCTGGATGGTTTGCCTCATAAACTGAAGGACATGTGGCGAAATGCGACCTCTCAATAAATTTGAAGATGATCGAATCCTCCATTCTAACTAATTCATCTCTAATATTTTGTAGATTTAAAACAGTTTCTGGTTTTGTGAAATCCATagacttcgtCCGCTTGTAATTAAAACTTAGATTAGATTGCTATGCTTTCTTTCTAATGAGCAAGAAGTAAAAAAAGTTGTAATAGAACAAGAAAAATGAAACTGAAACTTGAGAAATTGAAGACCGTTTATTAACTTAAATATCAATGGGAGGTCATCGAAAGAGAAAAAAATCAAAAAAAAAAATTTTCAAGAAAAAGAAACGTGATAAAAATTTTTATTGCCTTTTTCGACGAAGAAAAAGAAACGAGGCGGTCTCTTTTTTCTTTTCCAAACCTTTAGTACGGGTAATTAACGACACCCTAGAGGAAGAAAGAGGGGAAATTTAGTATGCTGTGCTTGGGTGTTTTGAAGTGGTACGGCGATGCGCGGAGTCCGAGAAAATCTGGAAGAGTAAAAAAGGAGTAGAAACATTTTGAAGCTATGGTGTGTGCGGCCGGCCTGGAAGTACCTTCAAAGAATGGGGTCTTATCTTGTTTTGCAAGTACCACTGAGCAGGATAATAATAGAAATGATAATATACTATAGTAGAGATAACGTCGATGACTTCCCATACTGTAATTGCTTTTAGTTGTGTATTTTTAGTGTGCAAGTTTCTGTAAATCGATTAATTTTTTTTTCTTTCCTCTTTTTATTAACCTTAATTTTTATTTTAGATTCCTGACTTCAACTCAAGACGCACAGATATTATAACATCTGCATAATAGGCATTTGCAAGAATTACTCGTGAGTAAGGAAAGAGTGAGGAACTATCGCATACCTGCATTTAAAGATGCCGATTTGGGCGCGAATCCTTTATTTTGGCTTCACCCTCATACTATTATCAGGGCCAGAAAAAGGAAGTGTTTCCCTCCTTCTTGAATTGATGTTACCCTCATAAAGCACGTGGCCTCTTATCGAGAAAGAAATTACCGTCGCTCGTGATTTGTTTGCAAAAAGAACAAAACTGAAAAAACCCAGACACGCTCGACTTCCTGTCTTCCTATTGATTGCAGCTTCCAATTTCGTCACACAACAAGGTCCTAGCGACGGCTCACAGGTTTTGTAACAAGCAATCGAAGGTTCTGGAATGGCGGGAAAGGGTTTAGTACCACATGCTATGATGCCCACTGTGATCTCCAGAGCAAAGTTCGTTCGATCGTACTGTTACTCTCTCTCTTTCAAACAGAATTGTCCGAATCGTGTGACAACAACAGCCTGTTCTCACACACTCTTTTCTTCTAACCAAGGGGGTGGTTTAGTTTAGTAGAACCTCGTGAAACTTACATTTACATATATATAAACTTGCATAAATTGGTCAATGCAAGAAATACATATTTGGTCTTTTCTAATTCGTAGTTTTTCAAGTTCTTAGATGCTTTCTTTTTCTCTTTTTTACAGATCATCAAGGAAGTAATTATCTACTTTTTACAACAAATATAAAACAAGGATCCaagtctactATGAGTGAATCTCCAATGTTCGCTGCCAACGGCATGCCAAAGGTAAATCAAGGTGCTGAAGAAGATGTCAGAATTTTAGGTTACGACCCATTAGCTTCTCCAGCTCTCCTTCAAGTGCAAATCCCAGCCACACCAACTTCTTTGGAAACTGCCAAGAGAGGTAGAAGAGAAGCTATAGATATTATTACCGGTAAAGACGACAGAGTTCTTGTCATTGTCGGTCCTTGTTCCATCCATGATCTAGAAGCCGCTCAAGAATACGCTTTGAGATTAAAGAAATTGTCAGATGAATTAAAAGGTGATTTATCCATCATTATGAGAGCATACTTGGAGAAGCCAAGAACAACCGTCGGCTGGAAAGGTCTAATTAATGACCCTGATGTTAACAACACTTTCAACATCAACAAGGGTTTGCAATCCGCTAGACAATTGTTTGTCAACTTGACAAATATCGGTTTGCCAATTGGTTCTGAAATGCTTGATACCATTTCTCCTCAATACTTGGCTGATTTGGTCTCCTTCGGTGCCATTGGTGCCAGAACCACCGAATCTCAACTGCACAGAGAATTGGCCTCCGGTTTGTCTTTCCCAGTTGGTTTCAAGAACGGTACCGATGGTACCTTAAATGTTGCTGTGGATGCTTGTCAAGCCGCTGCTCATTCTCACCATTTCATGGGTGTTACTTTACATGGTGTTGCTGCTATCACCACTACTAAGGGTAACGAACACTGCTTCGTTATTCTAAGAGGTGGTAAAAAGGGTACCAACTACGACGCTAAGTCCGTTGCAGAAGCTAAGGCTCAATTGCCTGCCGGTTCCAACGGTCTAATGATTGACTACTCTCACGGTAACTCCAATAAGGATTTCAGAAACCAACCAAAGGTCAATGACGTTGTTTGTGAGCAAATCGCTAACGGTGAAAACGCCATTACCGGTGTCATGATTGAATCAAACATCAACGAAGGTAACCAAGGCATCCCAGCCGAAGGTAAAGCCGGCTTGAAATATGGTGTTTCCATCACTGATGCTTGTATAGGTTGGGAAACTACTGAAGACGTCTTGAGGAAATTGGCTGCTGCTGTCAGACAAAGAAGAGAAGTTAACAAGAAATAGATTCTGCAGGTAGGGAAAGATATGAGCTATACAGCGGAATTTCCATATCACTCAGATTTTGTTATCTAATTTTTTCCTTCCCACGTCCGCGGGAATCTGTGTATATTACTGCATCTAGATATATGTTATCTTATCTTGGCGCGTACATTTAATTTTCAACGTATTCTATAAGAAATTGCGGGAGTTTTTTTCATGTAGATGATACTGACTGCACGCAAATATAGGCATGATTTATAGGCATGATTTGATGGCTGTACCGATAGGAACGCTAAGAGTAACTTCAGAATCGTTATCCTGGCGGAAAAAATTCATTTGTAAACTTTAAAAAAAAAAGCCAATATCCCCAAAATTATTAAGAGCGCCTCCATTATTAACTAAAATTTCACTCAGCATCCACAATGTATCAGGTATCTACTACAGATATTACATGTGGCGAAAAAGACAAGAACAATGCAATAGCGCATCAAGAAAAAACACAAAGCTTTCAATCAATGAATCGAAAATGTCATTAAAATAGTATATAAATTGAAACTAAGTCATAAAGCTATAAAAAGAAAATTTATTTAAATGCAAGATTTAAAGTAAATTCACTTAGGATCCGTATCTTGGAATTGGGATCAATTTTAAAGGTTTAGTTTTCTGCAAGGCTATACCAAACTTTTCATCCATGTCCAAATCTTTAGGTGAAATGTCTCCCTCTAATTTCCAGTTGAAGAATTGCAATAATGTGGCCAACATCAATGTCAACATCCTTATGGCCAAATTCATACCAGGGCAAATTCTTCTACCAGCACCAAAAGGCAATAAACCAAAATCTCTTCCCTTTACATCAATCTCACAACCTATAAATCTTTCAGGTGAGAAAATATCGGCATTTTGCCAAGCATTTGGATCTCTACCTATGGCCCACAAGTTTACCAAAATTTGAGCATCCTTTGGAACAATGTATCCGTACAATTCTACGTCTGTGTCGGCTTTTCTAGGCAACAAGAAAACTGTAGGTGGGTGCAACCTCAAAGTTTCCTTAATGATTGCTTGCAAGTATGGCAAATTTATTATGTCTGACTCTTGGATTTGCTTATCCTTACCCAATACCTGTTTTATTTCCTCTTGGGCCTTTTCCATCATTTCAGGGTTTCTTATTAACTCAGTCATAACCCACTCAAAGGTAGATGATGTAGTATCAGTACCGGCATCGAAAATATCGACTAACAAGTGGTTAATCTCTCCCATAGTTAATTCATTCTGCTTAAACAATTGCAATAATACGTCCAAGACATCATCTGTAGTTGTAGTAGTTGTAGTTGATGAATCTGGAGCTAACCTCTCACATATTATTCCTTGAAATACTGCAATTAACTTATCAAAAGAACATGCCAACCTTCTTCTAATTCCTGATGGATCGACGCAACCTAAAATTGGGAAGTAATCGGCGTAGTTTGGTTTACCGATGTCTTCCATAATATTCCAAATTAATTCCTTAAACTCTTGAGAGGTATGTGATTTATGATGGGCCAATTCAACAGAAAAGAATAATTTAGACAATAAATTCAAAGAAGTTGTGAAGGCAGCTTTACCAATGTCAACTGCCTGACCCTTCTGAGCGCACTCTTGAACGTATTCATATAATTGCTGAACCTTTGCATGTCTAAAGGTCTGGCATGCATCTAATCTTTGTGGAGACAATAAATGAACAGCAGTTATCTTTCTGAAGTTTCTCCATTTAGGTGAAACAGGCAACCAAGACATAGTCAATTTGTGGTGGTCACCAGCAGTTACTGAATTTGGAATTGTCCTGTTTGATAAAGGGTGATCCTTCTTCAAGAACATCTCTTTGGCGACATCAGCAGATGATACAACAATGGTGGTAACTGATCCTAACCTCAAAGAGATTAAAGGACCGTGGATCTTAGCCAAATTTGCAAATGATCTATGAGGCTTCTTTCCTACTTCCAAGATATTACCAATAATTGGTAAAGGTTTAGGACCTGGAGGTAACAATTTTGTTGTTTGTTGTGAGAATAATAATTTAATAAAATGAAATGAAATAAACCAAATTGCTAATATCATAGCTAATGTTGCATGATCCATTTTTAGTTTATGTATGTGTTTTTTGTAGTTATAGATTTAAGCAAGAAAAGAATACAAACAAAAAATTGAAAAAGATTGATTTAGAATTAAAAAGAAAAATATTTACGTAAGAAGGGAAAATAGTAAATGTTGCAAGTTCACTAAACTCCTAAATTATGCTGCCCTTTATATTCCCTGTTACAGCAGCCGAGCCAAAGGTATATAGGCTCCTTTGCATTAGCATGCGTAACAAACCACCTGTCAGTTTCAACCGAGGTGGTATCCGAGAGAATTGTGTGATTGCTTTAATTAATTTCGGAGAATCTCACATGCCACTGAAGATTAAAAACTGGATGCCAGAAAAGGGGTGTCCAGGTGTAACATCAATAGAGGAAGCTGAAAAGTCTTAGAACGGGTAATCTTCCACCAACCTGATGGGTTCCTAGATATAATCTCGAAGGGAATAAGTAGGGTGATACCGCAGAAGTGTCTGAATGTATTAAGGTCCTCACAGTTTAAATCCCGCTCACACTAACGTAGGATTATTATAACTCAAAAAAATGGCATTATTCTAAGTAAGTTAAATATCCGTAATCTTTAAACAagacttcgTCACCCATGAACCACACGGTTAGTCCAAAAGGGGCAGTTCAGATTCCAGATGCGGGAATTAGCTTGCTGCCACCCTCACCTCACTAACGCTGCGGTGTGCGGATACTTCATGCTATTTATAGACGCGCGTGTCGGAATCAGCACGCGCAAGAACCAAATGGGAAAATCGGAATGGGTCCAGAACTGCTTTGAGTGCTGGCTATTGGCGTCTGATTTCCGTTTTGGGAATCCTTTGCCGCGCGCCCCTCTCAAAACTCCGCACAAGTCCCAGAAAGCGGGAAAGAAATAAAACGCCACCAAAAAAAAAAAAATAAAAGCCAATCCTCGAAGCGTGGGTGGTAGGCCCTGGATTATCCCGTACAAGTATTTCTCAGGAGTAAAAAAACCGTTTGTTTTGGAATTTCCCATTTCGCGGCCACCTACGCCGCTATCTTTGCAACAACTATCTGCGATAACTCAGCAAATTTTGCATATTCGTGTTGCAGTATTGCGATAATGGGAGTCTTACTTCCAACATAACGGCAGAAAGAAATGTGAGAAAATTTTGCATCCTTTGCCTCCGTTCAAGTATATAAAGTCGGCATGCTTGATAATCTTTCTTTCCATCCTACATTGTTCTAATTATTCTTATTCTCCTTTATTCTTTCCTAACATACCAAGAAATTAATCTTCTGTCATTCGCTTAAACACTATATCAATAAAGATCTAAAAATGAAGGGAACCTACTACATCAACCACGGTGACCCATTGATGTACTTGAAGAAGCACATCAAGTTGAGACAATTCTTGGAAGGTTGGCAAGAAAACGTTGTTATCGAAAAGCCAAAGTCTATCTTGATCATCAGTGCACACTGGGACACCAACGTTCCAACCGTTAACTTCGTTGAACACTGTGACACCATCCACGACTTCGACGACTACCCAGACCCATTGTACCAAATCCAATACCGCGCTCCAGGTGCTCCAAACTTGGCTAAGAAGGTTGAAGAATTGCTTAAGGAATCTGGTATGGAATGTGAAATCGACACCAAGAGAGGTTTGGACCACGCTGCTTGGTTCCCATTGATGTTCATGTACCCAGAAGCTAACATCCCAATCTGTGAATTGTCTGTTCAACCATCTAAGGACGGTATCCACCACTACAACGTTGGTAAAGCTTTGTCTCCATTGTTGCAACAAGGTGTTTTGATCATCGGTTCTGGTGGAACCGTTCACCCATCTGACGACACCCCACACTGTCCAAACGGTGTTGCTCCATGGGCTATCGAGTTCGACAACTGGTTGGAAGACGCTTTGCTCTCGGGTAGATACGAAGACGTTAACAACTTCAAGAAGTTGGCTCCAAACTGGGAAATCTCTCACCCAGGTCAAGAACACTTGTACCCATTGCACGTTGCTTTGGGTGCTGCTGGTAAGAACCCAAAGACCCAATTGATCCACCGGTCTTGGGCTGCTAACGGTGTTTTCGGTTACTCTACCTACAACTTCACCCCAACCACCCAAAAGACCGACGGATCCTAAATCGCGTGCATTCATCCGCTCTAACCGAAAAGGAAGGAGTTAGACAACCTGAAGTCTAGGTCCCTATTTATTTTTTTATAGTTATGTTAGTATTAAGAACGTTATTTATATTTCAAATTTTTCTTTTTTTTCTGTACAGACGCGTGTACGCATGTAACATTATACTGAAAACCTTGCTTGAGAAGGTTTTGGGACGCTCGAAGATCGCGTCAGCTGAAGCTTCGTACGCTGCAGGTCGACAATTCTGCAGGTCGACAACCCTTAATATAACTTCGTATAATGTATGCTATACGAAGTTATTAGGTCTAGAGATCTGTTTAGCTTGCCTCGTCCCCGCCGGGTCACCCGGCCAGCGACATGGAGGCCCAGAATACCCTCCTTGACAGTCTTGACGTGCGCAGCTCAGGGGCATGATGTGACTGTCGCCCGTACATTTAGCCCATACATCCCCATGTATAATCATTTGCATCCATACATTTTGATGGCCGCACGGCGCGAAGCAAAAATTACGGCTCCTCGCTGCAGACCTGCGAGCAGGGAAACGCTCCCCTCACAGACGCGTTGAATTGTCCCCACGCCGCGCCCCTGTAGAGAAATATAAAAGGTTAGGATTTGCCACTGAGGTTCTTCTTTCATATACTTCCTTTTAAAATCTTGCTAGGATACAGTTCTCACATCACATCCGAACATAAACAACCATGGGTACCACTCTTGACGACACGGCTTACCGGTACCGCACCAGTGTCCCGGGGGACGCCGAGGCCATCGAGGCACTGGATGGGTCCTTCACCACCGACACCGTCTTCCGCGTCACCGCCACCGGGGACGGCTTCACCCTGCGGGAGGTGCCGGTGGACCCGCCCCTGACCAAGGTGTTCCCCGACGACGAATCGGACGACGAATCGGACGACGGGGAGGACGGCGACCCGGACTCCCGGACGTTCGTCGCGTACGGGGACGACGGCGACCTGGCGGGCTTCGTGGTCGTCTCGTACTCCGGCTGGAACCGCCGGCTGACCGTCGAGGACATCGAGGTCGCCCCGGAGCACCGGGGGCACGGGGTCGGGCGCGCGTTGATGGGGCTCGCGACGGAGTTCGCCCGCGAGCGGGGCGCCGGGCACCTCTGGCTGGAGGTCACCAACGTCAACGCACCGGCGATCCACGCGTACCGGCGGATGGGGTTCACCCTCTGCGGCCTGGACACCGCCCTGTACGACGGCACCGCCTCGGACGGCGAGCAGGCGCTCTACATGAGCATGCCCTGCCCCTAATCAGTACTGACAATAAAAAGATTCTTGTTTTCAAGAACTTGTCATTTGTATAGTTTTTTTATATTGTAGTTGTTCTATTTTAATCAAATGTTAGCGTGATTTATATTTTTTTTCGCCTCGACATCATCTGCCCAGATGCGAAGTTAAGTGCGCAGAAAGTAATATCATGCGTCAATCGTATGTGAATGCTGGTCGCTATACTGCTGTCGATTCGATACTAACGCCGCCATCCAGTGTCGAAAACGAGCTCTCGAGAACCCTTAAT'\n",
    "BTX['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "BTX['Primer_up_U']='GV1' #user site up\n",
    "BTX['Primer_down_U']='GV2'#user site down\n",
    "BTX['Primer_up']=''\n",
    "BTX['Primer_up_Tm']=''\n",
    "BTX['Primer_down']=''\n",
    "BTX['Primer_down_Tm']=''\n",
    "\n",
    "\n",
    "## CDSs ups and downs regions will be linked to KanMx to generate KO plasmids\n",
    "KanMX={}\n",
    "KanMX['name']='pTEF::KanMX::tTef'\n",
    "KanMX['seq']='TAGGTCTAGAGATCTGTTTAGCttgcctcgtccccgccgggtcacccggccagcgacatggaggcccagaataccctccttgacagtcttgacgtgcgcagctcaggggcatgatgtgactgtcgcccgtacatttagcccatacatccccatgtataatcatttgcatccatacattttgatggccgcacggcgcgaagcaaaaattacggctcctcgctgcagacctgcgagcagggaaacgctcccctcacagacgcgttgaattgtccccacgccgcgcccctgtagagaaatataaaaggttaggatttgccactgaggttcttctttcatatacttccttttaaaatcttgctaggatacagttctcacatcacatccgaacataaacaaccatgggtaaggaaaagactcacgtttcgaggccgcgattaaattccaacatggatgctgatttatatgggtataaatgggctcgcgataatgtcgggcaatcaggtgcgacaatctatcgattgtatgggaagcccgatgcgccagagttgtttctgaaacatggcaaaggtagcgttgccaatgatgttacagatgagatggtcagactaaactggctgacggaatttatgcctcttccgaccatcaagcattttatccgtactcctgatgatgcatggttactcaccactgcgatccccggcaaaacagcattccaggtattagaagaatatcctgattcaggtgaaaatattgttgatgcgctggcagtgttcctgcgccggttgcattcgattcctgtttgtaattgtccttttaacagcgatcgcgtatttcgtctcgctcaggcgcaatcacgaatgaataacggtttggttgatgcgagtgattttgatgacgagcgtaatggctggcctgttgaacaagtctggaaagaaatgcataagcttttgccattctcaccggattcagtcgtcactcatggtgatttctcacttgataaccttatttttgacgaggggaaattaataggttgtattgatgttggacgagtcggaatcgcagaccgataccaggatcttgccatcctatggaactgcctcggtgagttttctccttcattacagaaacggctttttcaaaaatatggtattgataatcctgatatgaataaattgcagtttcatttgatgctcgatgagtttttctaatcagtactgacaataaaaagattcttgttttcaagaacttgtcatttgtatagtttttttatattgtagttgttctattttaatcaaatgttagcgtgatttatattttttttcgcctcgacatcatctgcccagatgcgaagttaagtgcgcagaaagtaatatcatgcgtcaatcgtatgtgaatgctggtcgctatactgctgtcgattcgatactaacgccgccatccagtgtcgaaaacgaGCTCTCGAGAACCCTTAAT'\n",
    "KanMX['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "KanMX['Primer_up_U']='U1' #user site up\n",
    "KanMX['Primer_down_U']='U2'#user site down\n",
    "KanMX['Primer_up']=''\n",
    "KanMX['Primer_up_Tm']=''\n",
    "KanMX['Primer_down']=''\n",
    "KanMX['Primer_down_Tm']=''\n",
    "\n",
    "\n",
    "\n",
    "##in addition,CDS will be PCRed with the forward primer of the Up and the reverse of the down\n",
    "#and annealed into pCCM022 to be flanked by  HomoB and XII-dwn\n",
    "\n",
    "KGD2_up={}\n",
    "KGD2_up['name']='KGD2_up'\n",
    "KGD2_up['seq']='ggcaggaccttagttttgattggctgcaaaaaaagggttgttaatatcgtctttgacggagatgcgggtggctggaattgttttgtttattattccccaagagagaagagacaagaagtagtaatgacgatggaacaaagatcggttctaataggaccacagggttgaatagagacgagtaaagacgcaacggtcaaggctttccacgagacgttcgaat'\n",
    "KGD2_up['need_reverse']=1 #if parts needs to be reversed:1 else:0\n",
    "KGD2_up['Primer_up_U']='GV1' #user site up\n",
    "KGD2_up['Primer_down_U']='U1'#user site down\n",
    "KGD2_up['Primer_up']=''\n",
    "KGD2_up['Primer_up_Tm']=''\n",
    "KGD2_up['Primer_down']=''\n",
    "KGD2_up['Primer_down_Tm']=''\n",
    "\n",
    "KGD2_dw={}\n",
    "KGD2_dw['name']='KGD2_dw'\n",
    "KGD2_dw['seq']='atgtggaaatcaaaagaatattagttgattctattatataataaatagttataatcattgtattttgatgatgaaaattcagtatctaagagttatagccaagagagtaatttttaatacttttttgcgcttttccatatctcattataaaaagaaaacattaattcatagttccatttatcatcatagcatatctttaa'\n",
    "KGD2_dw['need_reverse']=1 #if parts needs to be reversed:1 else:0\n",
    "KGD2_dw['Primer_up_U']='U2' #user site up\n",
    "KGD2_dw['Primer_down_U']='GV2'#user site down\n",
    "KGD2_dw['Primer_up']=''\n",
    "KGD2_dw['Primer_up_Tm']=''\n",
    "KGD2_dw['Primer_down']=''\n",
    "KGD2_dw['Primer_down_Tm']=''\n",
    "\n",
    "KGD2=[KGD2_up,KanMX,KGD2_dw]\n",
    "\n",
    "HEM25_up={}\n",
    "HEM25_up['name']='HEM25_up'\n",
    "HEM25_up['seq']='atttcacttcgtccttaatattttaacacattaactgtttgctataagcatacattgcttttaccactttttttaataatatctttttatccgtattaattaccgcatataattttagttcgaaacattaaaggtcagtcccgaagttataattataacatttaattcctgaatttaccgtattacatttaaatttgcat'\n",
    "HEM25_up['need_reverse']=1 #if parts needs to be reversed:1 else:0\n",
    "HEM25_up['Primer_up_U']='GV1' #user site up\n",
    "HEM25_up['Primer_down_U']='U1'#user site down\n",
    "HEM25_up['Primer_up']=''\n",
    "HEM25_up['Primer_up_Tm']=''\n",
    "HEM25_up['Primer_down']=''\n",
    "HEM25_up['Primer_down_Tm']=''\n",
    "\n",
    "HEM25_dw={}\n",
    "HEM25_dw['name']='HEM25_dw'\n",
    "HEM25_dw['seq']='gtgcacttatccatcgactctcttatatttatttaaacacttgtaacgcatccaatctatttcatagttagagtctctcttccttgttattgatatgatttttcttttgtaccgtgtgtgtatgtatgtatgtatgaacgtactactaagcttcaacgtcactctataaagaaactaaaaagacacgtatttatacctt'\n",
    "HEM25_dw['need_reverse']=1 #if parts needs to be reversed:1 else:0\n",
    "HEM25_dw['Primer_up_U']='U2' #user site up\n",
    "HEM25_dw['Primer_down_U']='GV2'#user site down\n",
    "HEM25_dw['Primer_up']=''\n",
    "HEM25_dw['Primer_up_Tm']=''\n",
    "HEM25_dw['Primer_down']=''\n",
    "HEM25_dw['Primer_down_Tm']=''\n",
    "\n",
    "\n",
    "HEM25=[HEM25_up,KanMX,HEM25_dw]\n",
    "\n",
    "\n",
    "\n",
    "VMA16_up={} #PPA1\n",
    "VMA16_up['name']='VMA16_up'\n",
    "VMA16_up['seq']='ggcctattgggtatctgcttgtcaggtgcaggcccaactatcttggctttggccactgagaatttcgaagaaatctctcaagaaattatcaacaggttcgccaaaaacggcatcaagtgctcctggaaactactggagcctgcctacgatggtgctagcgtcgaacagcaatgaaatgtcctttacccctagtatactgtgtt'\n",
    "VMA16_up['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "VMA16_up['Primer_up_U']='GV1' #user site up\n",
    "VMA16_up['Primer_down_U']='U1'#user site down\n",
    "VMA16_up['Primer_up']=''\n",
    "VMA16_up['Primer_up_Tm']=''\n",
    "VMA16_up['Primer_down']=''\n",
    "VMA16_up['Primer_down_Tm']=''\n",
    "\n",
    "VMA16_dw={}\n",
    "VMA16_dw['name']='VMA16_dw'\n",
    "VMA16_dw['seq']='agcgcatccatatttactaatagatataatgagatatctgagatacatgtgtatgtatatatgcacgttttcttttattatctaaaaatcatattatattaagtaagagaaaaaaatgtacaactatataaatatatatttatttaaaatggttttgaatttttcctattctggttgatattgcccaaaagctattcagt'\n",
    "VMA16_dw['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "VMA16_dw['Primer_up_U']='U2' #user site up\n",
    "VMA16_dw['Primer_down_U']='GV2'#user site down\n",
    "VMA16_dw['Primer_up']=''\n",
    "VMA16_dw['Primer_up_Tm']=''\n",
    "VMA16_dw['Primer_down']=''\n",
    "VMA16_dw['Primer_down_Tm']=''\n",
    "VMA16 = [VMA16_up,KanMX,VMA16_dw]\n",
    "\n",
    "\n",
    "\n",
    "\n",
    "CAX4_up={} \n",
    "CAX4_up['name']='CAX4_up'\n",
    "CAX4_up['seq']='aaccccgagaatttaaatagaacagagtatatgcatttattaacagttattacgccttgtccccagggagagtagtaagggatatgggcatttataaatgtacataagtcacgtttttgcagccaataaaatataataatagaatagtaacaaggaaagctaggccaaaactccttacatggagctagtatacccctttt'\n",
    "CAX4_up['need_reverse']=1 #if parts needs to be reversed:1 else:0\n",
    "CAX4_up['Primer_up_U']='GV1' #user site up\n",
    "CAX4_up['Primer_down_U']='U1'#user site down\n",
    "CAX4_up['Primer_up']=''\n",
    "CAX4_up['Primer_up_Tm']=''\n",
    "CAX4_up['Primer_down']=''\n",
    "CAX4_up['Primer_down_Tm']=''\n",
    "\n",
    "CAX4_dw={}\n",
    "CAX4_dw['name']='CAX4_dw'\n",
    "CAX4_dw['seq']='aagtgtgggtgacatgctaaatccgcggctttccttaagctttttcacaaaccatctattatgtccagaagctccgctgattatccgtggaaaagggtcttattctgccgtaggaaacttcttctcgagaaaaaagtgagttcacccgcggcgctctcgtaaaacccgagggccgattctttccctccgtggaacaatcg'\n",
    "CAX4_dw['need_reverse']=1 #if parts needs to be reversed:1 else:0\n",
    "CAX4_dw['Primer_up_U']='U2' #user site up\n",
    "CAX4_dw['Primer_down_U']='GV2'#user site down\n",
    "CAX4_dw['Primer_up']=''\n",
    "CAX4_dw['Primer_up_Tm']=''\n",
    "CAX4_dw['Primer_down']=''\n",
    "CAX4_dw['Primer_down_Tm']=''\n",
    "CAX4 = [CAX4_up,KanMX,CAX4_dw]\n",
    "\n",
    "\n",
    "PRO2_up={} #PPA1\n",
    "PRO2_up['name']='PRO2_up'\n",
    "PRO2_up['seq']='ttgtaaagtgaccttgttttgactttttaacgagtaaaaaaacgagttaacaaattcttgacgtaaacgataaagccgcccatcaaaagaaatgcaataatgtcaatgatgaaatacctatacaggtagatggatttcgtgttgttcttcgatttatttaacacgtttttcagctctatcaaactatccatttcccttag'\n",
    "PRO2_up['need_reverse']=1 #if parts needs to be reversed:1 else:0\n",
    "PRO2_up['Primer_up_U']='GV1' #user site up\n",
    "PRO2_up['Primer_down_U']='U1'#user site down\n",
    "PRO2_up['Primer_up']=''\n",
    "PRO2_up['Primer_up_Tm']=''\n",
    "PRO2_up['Primer_down']=''\n",
    "PRO2_up['Primer_down_Tm']=''\n",
    "\n",
    "PRO2_dw={}\n",
    "PRO2_dw['name']='PRO2_dw'\n",
    "PRO2_dw['seq']='gagtagatgtaagacgtgaaaatgccatggtaaaacgtattgtttagtgaaatgtatactaagtaggagtaaaaatcagcaatttctggtgttgaaatgtgaacaacagcgttctcctccgaaaaagcaagctataacgttgctcaacttttgattccgaaatcagaagcttaaataaggaaaactgaaagctaattgga'\n",
    "PRO2_dw['need_reverse']=1 #if parts needs to be reversed:1 else:0\n",
    "PRO2_dw['Primer_up_U']='U2' #user site up\n",
    "PRO2_dw['Primer_down_U']='GV2'#user site down\n",
    "PRO2_dw['Primer_up']=''\n",
    "PRO2_dw['Primer_up_Tm']=''\n",
    "PRO2_dw['Primer_down']=''\n",
    "PRO2_dw['Primer_down_Tm']=''\n",
    "PRO2 = [PRO2_up,KanMX,PRO2_dw]\n",
    "\n",
    "\n",
    "GLR1_up={} #PPA1\n",
    "GLR1_up['name']='GLR1_up'\n",
    "GLR1_up['seq']='tggtcaattctatgccttttatgcacattgcatgagtattctaaaaagatgctgcatgctgcccgtaaatcttcattattttcagagtcaggtacggaaaagacgacagtttctccgtataacagcattgaaagcgtggaagaatcaaactcggctctagattttacgcgtttagcataaaattgttggttttatacgta'\n",
    "GLR1_up['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "GLR1_up['Primer_up_U']='GV1' #user site up\n",
    "GLR1_up['Primer_down_U']='U1'#user site down\n",
    "GLR1_up['Primer_up']=''\n",
    "GLR1_up['Primer_up_Tm']=''\n",
    "GLR1_up['Primer_down']=''\n",
    "GLR1_up['Primer_down_Tm']=''\n",
    "\n",
    "GLR1_dw={}\n",
    "GLR1_dw['name']='GLR1_dw'\n",
    "GLR1_dw['seq']='cttttactgctttctaactttaagccttcaaagaagaagctcttctctttctgatttcagccttctcagcctttctttcagacaatctcttagccaacaattgagcgtattcggcagcagcttctctttgagcttgagcgtttctgaccttcaaagctctttggtgtctctttctttgcaatctttgaggagtaaccaatc'\n",
    "GLR1_dw['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "GLR1_dw['Primer_up_U']='U2' #user site up\n",
    "GLR1_dw['Primer_down_U']='GV2'#user site down\n",
    "GLR1_dw['Primer_up']=''\n",
    "GLR1_dw['Primer_up_Tm']=''\n",
    "GLR1_dw['Primer_down']=''\n",
    "GLR1_dw['Primer_down_Tm']=''\n",
    "GLR1 = [GLR1_up,KanMX,GLR1_dw]\n",
    "\n",
    "\n",
    "YSY6_up={} \n",
    "YSY6_up['name']='YSY6_up'\n",
    "YSY6_up['seq']='ggagtcttcttgcgagaaaagaaataattataatataaaaaaaaattgaatggagataagaagtttaactagcaaagttcgagagatataaaaagtagagatgcgaaaataaaattaaaacaaatctgaaaataatggaagtgaaacaaggctaagcaagaaaattttctatattgccaagacaggaacgtttgtgatga'\n",
    "YSY6_up['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "YSY6_up['Primer_up_U']='GV1' #user site up\n",
    "YSY6_up['Primer_down_U']='U1'#user site down\n",
    "YSY6_up['Primer_up']=''\n",
    "YSY6_up['Primer_up_Tm']=''\n",
    "YSY6_up['Primer_down']=''\n",
    "YSY6_up['Primer_down_Tm']=''\n",
    "\n",
    "YSY6_dw={}\n",
    "YSY6_dw['name']='YSY6_dw'\n",
    "YSY6_dw['seq']='gcagtattacccgcgttgtcatttatggatgcgaaaaaagatgaagtaattcaagtgctattgtgatttaccatgccaatcaaacgtgcaacaaagatgagtatgaggggggacccaggcatagtacacagaaaagctttgtagactgtctttgttttcttaatttttcaagtaggctttctgcttttcttctgtactta'\n",
    "YSY6_dw['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "YSY6_dw['Primer_up_U']='U2' #user site up\n",
    "YSY6_dw['Primer_down_U']='GV2'#user site down\n",
    "YSY6_dw['Primer_up']=''\n",
    "YSY6_dw['Primer_up_Tm']=''\n",
    "YSY6_dw['Primer_down']=''\n",
    "YSY6_dw['Primer_down_Tm']=''\n",
    "YSY6 = [YSY6_up,KanMX,YSY6_dw]\n",
    "\n",
    "\n",
    "\n",
    "\n",
    "PDX3_up={} \n",
    "PDX3_up['name']='PDX3_up'\n",
    "PDX3_up['seq']='aggtagagtatctttaaaatatctaatcatgcaaattagtcatttcttctttcttataagttgcggccccttctagtatttacaatttacccggcgctggttctggtggcatctataagagctctatctggctgctaagcagacttgctgttacttgtatgtgctgtttatcaagattggggcattacgcatgtgtatgt'\n",
    "PDX3_up['need_reverse']=1 #if parts needs to be reversed:1 else:0\n",
    "PDX3_up['Primer_up_U']='GV1' #user site up\n",
    "PDX3_up['Primer_down_U']='U1'#user site down\n",
    "PDX3_up['Primer_up']=''\n",
    "PDX3_up['Primer_up_Tm']=''\n",
    "PDX3_up['Primer_down']=''\n",
    "PDX3_up['Primer_down_Tm']=''\n",
    "\n",
    "PDX3_dw={}\n",
    "PDX3_dw['name']='PDX3_dw'\n",
    "PDX3_dw['seq']='tctttggatgaatttgtcggttctaactcttttttggaaacggcaaggtgagcagtaggcagctcatctcttcaactatttttataacatataatttccctttgtgaaaaaattttcccgccatttccaaatttcggaattcctttttcttttttttcatctcggtatattctgacatattaccctataggcggtataaa'\n",
    "PDX3_dw['need_reverse']= 1 #if parts needs to be reversed:1 else:0\n",
    "PDX3_dw['Primer_up_U']='U2' #user site up\n",
    "PDX3_dw['Primer_down_U']='GV2'#user site down\n",
    "PDX3_dw['Primer_up']=''\n",
    "PDX3_dw['Primer_up_Tm']=''\n",
    "PDX3_dw['Primer_down']=''\n",
    "PDX3_dw['Primer_down_Tm']=''\n",
    "PDX3 = [PDX3_up,KanMX,PDX3_dw]"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 10,
   "metadata": {},
   "outputs": [],
   "source": [
    "UpXII5={}\n",
    "UpXII5['name']='UpXII5'\n",
    "UpXII5['seq']='GGGGGGGGGGGGGGGGGGGGGGGGGGGccggaggaggccgctataaccggtttgaatttattgtcacagtgtcacatcagc'\n",
    "UpXII5['need_reverse']= 0 #if parts needs to be reversed:1 else:0\n",
    "UpXII5['Primer_up_U']='U2' #user site up\n",
    "UpXII5['Primer_down_U']='GV1'#user site down\n",
    "UpXII5['Primer_up']=''\n",
    "UpXII5['Primer_up_Tm']=''\n",
    "UpXII5['Primer_down']=''\n",
    "UpXII5['Primer_down_Tm']=''\n",
    "\n",
    "HomoB={}\n",
    "HomoB['name']='HomoB'\n",
    "HomoB['seq']='ACTGAATACAGGCTAGTGAATGCTCTCGGTCTTGTACGTGTGCGATTGACGGGGGGGGGGGGGGGGGGGGGGGGGGG'\n",
    "HomoB['need_reverse']= 0 #if parts needs to be reversed:1 else:0\n",
    "HomoB['Primer_up_U']='GV1' #user site up\n",
    "HomoB['Primer_down_U']='U2'#user site down\n",
    "HomoB['Primer_up']=''\n",
    "HomoB['Primer_up_Tm']=''\n",
    "HomoB['Primer_down']=''\n",
    "HomoB['Primer_down_Tm']=''\n",
    "\n",
    "HomoA={}\n",
    "HomoA['name']='HomoA'\n",
    "HomoA['seq']='GGGGGGGGGGGGGGGGGGGGGGGGGGGGTACTCGGCTATGAGATTGAATACTCACGAAGGTAACGGTGGCACGACAAC'\n",
    "HomoA['need_reverse']= 0 #if parts needs to be reversed:1 else:0\n",
    "HomoA['Primer_up_U']='GV1' #user site up\n",
    "HomoA['Primer_down_U']='U2'#user site down\n",
    "HomoA['Primer_up']=''\n",
    "HomoA['Primer_up_Tm']=''\n",
    "HomoA['Primer_down']=''\n",
    "HomoA['Primer_down_Tm']=''\n",
    "\n",
    "\n",
    "DwXII5={}\n",
    "DwXII5['name']='DwXII5'\n",
    "DwXII5['seq']='gcaactcagaagtttgacagcaagcaagttcatcattcgaactagccttattgttttagttcGGGGGGGGGGGGGGGGGGGGGGGGGGGG'\n",
    "DwXII5['need_reverse']= 0 #if parts needs to be reversed:1 else:0\n",
    "DwXII5['Primer_up_U']='GV1' #user site up\n",
    "DwXII5['Primer_down_U']='U2'#user site down\n",
    "DwXII5['Primer_up']=''\n",
    "DwXII5['Primer_up_Tm']=''\n",
    "DwXII5['Primer_down']=''\n",
    "DwXII5['Primer_down_Tm']=''\n",
    "\n",
    "\n",
    "\n",
    "BB_PCR = [UpXII5,DwXII5,HomoA,HomoB]"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 12,
   "metadata": {},
   "outputs": [
    {
     "name": "stdout",
     "output_type": "stream",
     "text": [
      "Will use input Tm_target for design:  50\n",
      "[{'name': 'UpXII5', 'seq': 'GGGGGGGGGGGGGGGGGGGGGGGGGGGCCGGAGGAGGCCGCTATAACCGGTTTGAATTTATTGTCACAGTGTCACATCAGC', 'need_reverse': 0, 'Primer_up_U': 'U2', 'Primer_down_U': 'GV1', 'Primer_up': 'ACACAGGCuGGGGGGGGGGG', 'Primer_up_Tm': 55.40352299998773, 'Primer_down': 'ATCGCACuGCTGATGTGACACTGTGAC', 'Primer_down_Tm': 50.61273634503215}, {'name': 'DwXII5', 'seq': 'GCAACTCAGAAGTTTGACAGCAAGCAAGTTCATCATTCGAACTAGCCTTATTGTTTTAGTTCGGGGGGGGGGGGGGGGGGGGGGGGGGGG', 'need_reverse': 0, 'Primer_up_U': 'GV1', 'Primer_down_U': 'U2', 'Primer_up': 'CGTGCGAuGCAACTCAGAAGTTTGACAGC', 'Primer_up_Tm': 52.21562927265825, 'Primer_down': 'AGCCTGTGuCCCCCCCCCCC', 'Primer_down_Tm': 55.40352299998773}, {'name': 'HomoA', 'seq': 'GGGGGGGGGGGGGGGGGGGGGGGGGGGGTACTCGGCTATGAGATTGAATACTCACGAAGGTAACGGTGGCACGACAAC', 'need_reverse': 0, 'Primer_up_U': 'GV1', 'Primer_down_U': 'U2', 'Primer_up': 'CGTGCGAuGGGGGGGGGGG', 'Primer_up_Tm': 55.40352299998773, 'Primer_down': 'AGCCTGTGuGTTGTCGTGCCACCG', 'Primer_down_Tm': 51.13037814321416}, {'name': 'HomoB', 'seq': 'ACTGAATACAGGCTAGTGAATGCTCTCGGTCTTGTACGTGTGCGATTGACGGGGGGGGGGGGGGGGGGGGGGGGGGG', 'need_reverse': 0, 'Primer_up_U': 'GV1', 'Primer_down_U': 'U2', 'Primer_up': 'CGTGCGAuACTGAATACAGGCTAGTGAATG', 'Primer_up_Tm': 51.38995044586477, 'Primer_down': 'AGCCTGTGuCCCCCCCCCCC', 'Primer_down_Tm': 55.40352299998773}]\n"
     ]
    }
   ],
   "source": [
    "Parts = dt.User_Assebmly(BB_PCR, 50, User_Overhangs)\n",
    "print(Parts)"
   ]
  },
  {
   "cell_type": "markdown",
   "metadata": {},
   "source": [
    "### Design"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 13,
   "metadata": {},
   "outputs": [
    {
     "ename": "NameError",
     "evalue": "name 'YER084W_KO_BTX' is not defined",
     "output_type": "error",
     "traceback": [
      "\u001b[0;31m---------------------------------------------------------------------------\u001b[0m",
      "\u001b[0;31mNameError\u001b[0m                                 Traceback (most recent call last)",
      "\u001b[0;32m<ipython-input-13-1fe7a029b64a>\u001b[0m in \u001b[0;36m<module>\u001b[0;34m\u001b[0m\n\u001b[1;32m      1\u001b[0m \u001b[0mimportlib\u001b[0m\u001b[0;34m.\u001b[0m\u001b[0mreload\u001b[0m\u001b[0;34m(\u001b[0m\u001b[0mdt\u001b[0m\u001b[0;34m)\u001b[0m\u001b[0;34m\u001b[0m\u001b[0;34m\u001b[0m\u001b[0m\n\u001b[0;32m----> 2\u001b[0;31m \u001b[0mParts\u001b[0m \u001b[0;34m=\u001b[0m \u001b[0mdt\u001b[0m\u001b[0;34m.\u001b[0m\u001b[0mUser_Assebmly\u001b[0m\u001b[0;34m(\u001b[0m\u001b[0mYER084W_KO_BTX\u001b[0m\u001b[0;34m,\u001b[0m \u001b[0;36m50\u001b[0m\u001b[0;34m,\u001b[0m \u001b[0mUser_Overhangs\u001b[0m\u001b[0;34m)\u001b[0m\u001b[0;34m\u001b[0m\u001b[0;34m\u001b[0m\u001b[0m\n\u001b[0m\u001b[1;32m      3\u001b[0m \u001b[0mprint\u001b[0m\u001b[0;34m(\u001b[0m\u001b[0mParts\u001b[0m\u001b[0;34m)\u001b[0m\u001b[0;34m\u001b[0m\u001b[0;34m\u001b[0m\u001b[0m\n",
      "\u001b[0;31mNameError\u001b[0m: name 'YER084W_KO_BTX' is not defined"
     ]
    }
   ],
   "source": [
    "importlib.reload(dt)\n",
    "Parts = dt.User_Assebmly(YER084W_KO_BTX, 50, User_Overhangs)\n",
    "print(Parts)"
   ]
  },
  {
   "cell_type": "markdown",
   "metadata": {},
   "source": [
    "#### compile into a table"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 14,
   "metadata": {
    "scrolled": false
   },
   "outputs": [
    {
     "data": {
      "text/html": [
       "<div>\n",
       "<style scoped>\n",
       "    .dataframe tbody tr th:only-of-type {\n",
       "        vertical-align: middle;\n",
       "    }\n",
       "\n",
       "    .dataframe tbody tr th {\n",
       "        vertical-align: top;\n",
       "    }\n",
       "\n",
       "    .dataframe thead th {\n",
       "        text-align: right;\n",
       "    }\n",
       "</style>\n",
       "<table border=\"1\" class=\"dataframe\">\n",
       "  <thead>\n",
       "    <tr style=\"text-align: right;\">\n",
       "      <th></th>\n",
       "      <th>Seq</th>\n",
       "      <th>Dir</th>\n",
       "      <th>Tm (c)</th>\n",
       "      <th>User</th>\n",
       "      <th>Template</th>\n",
       "      <th>Template_Len</th>\n",
       "      <th>benchling</th>\n",
       "    </tr>\n",
       "    <tr>\n",
       "      <th>Primer Name</th>\n",
       "      <th></th>\n",
       "      <th></th>\n",
       "      <th></th>\n",
       "      <th></th>\n",
       "      <th></th>\n",
       "      <th></th>\n",
       "      <th></th>\n",
       "    </tr>\n",
       "  </thead>\n",
       "  <tbody>\n",
       "    <tr>\n",
       "      <th>BTX.1</th>\n",
       "      <td>ACACAGGCuGGGGGGGGGGG</td>\n",
       "      <td>fwd</td>\n",
       "      <td>55.4</td>\n",
       "      <td>U2</td>\n",
       "      <td>UpXII5</td>\n",
       "      <td>81</td>\n",
       "      <td>ACACAGGCtGGGGGGGGGGG</td>\n",
       "    </tr>\n",
       "    <tr>\n",
       "      <th>BTX.1</th>\n",
       "      <td>ATCGCACuGCTGATGTGACACTGTGAC</td>\n",
       "      <td>rvs</td>\n",
       "      <td>50.61</td>\n",
       "      <td>GV1</td>\n",
       "      <td>UpXII5</td>\n",
       "      <td>81</td>\n",
       "      <td>ATCGCACtGCTGATGTGACACTGTGAC</td>\n",
       "    </tr>\n",
       "    <tr>\n",
       "      <th>BTX.1</th>\n",
       "      <td>CGTGCGAuGCAACTCAGAAGTTTGACAGC</td>\n",
       "      <td>fwd</td>\n",
       "      <td>52.22</td>\n",
       "      <td>GV1</td>\n",
       "      <td>DwXII5</td>\n",
       "      <td>90</td>\n",
       "      <td>CGTGCGAtGCAACTCAGAAGTTTGACAGC</td>\n",
       "    </tr>\n",
       "    <tr>\n",
       "      <th>BTX.1</th>\n",
       "      <td>AGCCTGTGuCCCCCCCCCCC</td>\n",
       "      <td>rvs</td>\n",
       "      <td>55.4</td>\n",
       "      <td>U2</td>\n",
       "      <td>DwXII5</td>\n",
       "      <td>90</td>\n",
       "      <td>AGCCTGTGtCCCCCCCCCCC</td>\n",
       "    </tr>\n",
       "    <tr>\n",
       "      <th>BTX.1</th>\n",
       "      <td>CGTGCGAuGGGGGGGGGGG</td>\n",
       "      <td>fwd</td>\n",
       "      <td>55.4</td>\n",
       "      <td>GV1</td>\n",
       "      <td>HomoA</td>\n",
       "      <td>78</td>\n",
       "      <td>CGTGCGAtGGGGGGGGGGG</td>\n",
       "    </tr>\n",
       "    <tr>\n",
       "      <th>BTX.1</th>\n",
       "      <td>AGCCTGTGuGTTGTCGTGCCACCG</td>\n",
       "      <td>rvs</td>\n",
       "      <td>51.13</td>\n",
       "      <td>U2</td>\n",
       "      <td>HomoA</td>\n",
       "      <td>78</td>\n",
       "      <td>AGCCTGTGtGTTGTCGTGCCACCG</td>\n",
       "    </tr>\n",
       "    <tr>\n",
       "      <th>BTX.1</th>\n",
       "      <td>CGTGCGAuACTGAATACAGGCTAGTGAATG</td>\n",
       "      <td>fwd</td>\n",
       "      <td>51.39</td>\n",
       "      <td>GV1</td>\n",
       "      <td>HomoB</td>\n",
       "      <td>77</td>\n",
       "      <td>CGTGCGAtACTGAATACAGGCTAGTGAATG</td>\n",
       "    </tr>\n",
       "    <tr>\n",
       "      <th>BTX.1</th>\n",
       "      <td>AGCCTGTGuCCCCCCCCCCC</td>\n",
       "      <td>rvs</td>\n",
       "      <td>55.4</td>\n",
       "      <td>U2</td>\n",
       "      <td>HomoB</td>\n",
       "      <td>77</td>\n",
       "      <td>AGCCTGTGtCCCCCCCCCCC</td>\n",
       "    </tr>\n",
       "  </tbody>\n",
       "</table>\n",
       "</div>"
      ],
      "text/plain": [
       "                                        Seq  Dir Tm (c) User Template  \\\n",
       "Primer Name                                                             \n",
       "BTX.1                  ACACAGGCuGGGGGGGGGGG  fwd   55.4   U2   UpXII5   \n",
       "BTX.1           ATCGCACuGCTGATGTGACACTGTGAC  rvs  50.61  GV1   UpXII5   \n",
       "BTX.1         CGTGCGAuGCAACTCAGAAGTTTGACAGC  fwd  52.22  GV1   DwXII5   \n",
       "BTX.1                  AGCCTGTGuCCCCCCCCCCC  rvs   55.4   U2   DwXII5   \n",
       "BTX.1                   CGTGCGAuGGGGGGGGGGG  fwd   55.4  GV1    HomoA   \n",
       "BTX.1              AGCCTGTGuGTTGTCGTGCCACCG  rvs  51.13   U2    HomoA   \n",
       "BTX.1        CGTGCGAuACTGAATACAGGCTAGTGAATG  fwd  51.39  GV1    HomoB   \n",
       "BTX.1                  AGCCTGTGuCCCCCCCCCCC  rvs   55.4   U2    HomoB   \n",
       "\n",
       "            Template_Len                       benchling  \n",
       "Primer Name                                               \n",
       "BTX.1                 81            ACACAGGCtGGGGGGGGGGG  \n",
       "BTX.1                 81     ATCGCACtGCTGATGTGACACTGTGAC  \n",
       "BTX.1                 90   CGTGCGAtGCAACTCAGAAGTTTGACAGC  \n",
       "BTX.1                 90            AGCCTGTGtCCCCCCCCCCC  \n",
       "BTX.1                 78             CGTGCGAtGGGGGGGGGGG  \n",
       "BTX.1                 78        AGCCTGTGtGTTGTCGTGCCACCG  \n",
       "BTX.1                 77  CGTGCGAtACTGAATACAGGCTAGTGAATG  \n",
       "BTX.1                 77            AGCCTGTGtCCCCCCCCCCC  "
      ]
     },
     "execution_count": 14,
     "metadata": {},
     "output_type": "execute_result"
    }
   ],
   "source": [
    "\n",
    "\n",
    "\n",
    "primer_name=[]\n",
    "primer_dir=[]\n",
    "primer_tm=[]\n",
    "primer_OH=[]\n",
    "primer_template_name=[]\n",
    "primer_seq=[]\n",
    "primer_template_len=[]\n",
    "benchling=[]\n",
    "i=108\n",
    "for part in Parts:\n",
    "    #here we adapt the order with which the overhang names are appended to match the Overhang names to the inversion of sequences\n",
    "    if part['need_reverse']==0:\n",
    "        primer_OH.append(part['Primer_up' +'_U'])\n",
    "        primer_OH.append(part['Primer_down'+'_U'])\n",
    "    elif part['need_reverse']==1:\n",
    "        primer_OH.append(part['Primer_down'+'_U'])\n",
    "        primer_OH.append(part['Primer_up' +'_U'])\n",
    "        \n",
    "    for primer in ['Primer_up', 'Primer_down']:\n",
    "        primer_name.append('BTX.1')#.zfill fills primer number wiht leading 0\n",
    "        primer_seq.append(part[primer])\n",
    "        primer_tm.append(round(part[primer +'_Tm'],2))\n",
    "        primer_template_len.append(len(part['seq']))\n",
    "        primer_template_name.append(part['name'])\n",
    "        #primer_OH.append(part[primer +'_U'])\n",
    "        primer_dir.append('fwd'  if primer[7:] == 'up' else 'rvs' )\n",
    "        benchling.append(part[primer].replace('u','t'))\n",
    "        i+=1\n",
    "## need to fix the current overhang name bug (if seq needs inversion the OH name doenst match)\n",
    "        \n",
    "\n",
    "\n",
    "df = np.stack([primer_name, primer_seq,primer_dir, primer_tm,primer_OH, primer_template_name, primer_template_len, benchling] ).T\n",
    "Table=pd.DataFrame(df , columns=['Primer Name', 'Seq','Dir', 'Tm (c)','User', 'Template', 'Template_Len','benchling'])\n",
    "\n",
    "Table.set_index('Primer Name')"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 77,
   "metadata": {},
   "outputs": [
    {
     "name": "stdout",
     "output_type": "stream",
     "text": [
      "BTX.112\n",
      "BTX.113\n",
      "BTX.114\n",
      "BTX.115\n",
      "BTX.116\n",
      "BTX.117\n",
      "BTX.118\n",
      "BTX.119\n",
      "BTX.120\n",
      "BTX.121\n",
      "BTX.122\n",
      "BTX.123\n",
      "BTX.124\n",
      "BTX.125\n",
      "BTX.126\n",
      "BTX.127\n",
      "BTX.128\n",
      "BTX.129\n",
      "BTX.130\n",
      "BTX.131\n",
      "BTX.132\n",
      "BTX.133\n",
      "BTX.134\n",
      "BTX.135\n",
      "BTX.136\n",
      "BTX.137\n",
      "BTX.138\n",
      "BTX.139\n",
      "BTX.140\n",
      "BTX.141\n",
      "BTX.142\n",
      "BTX.143\n"
     ]
    }
   ],
   "source": [
    "for i in range(112,112+32):\n",
    "    print('BTX.'+str(i))"
   ]
  },
  {
   "cell_type": "markdown",
   "metadata": {},
   "source": [
    "### #design primers for simple insertion in User plasmid"
   ]
  },
  {
   "cell_type": "code",
   "execution_count": 60,
   "metadata": {},
   "outputs": [
    {
     "name": "stdout",
     "output_type": "stream",
     "text": [
      "Will use input Tm_target for design:  50\n",
      "[{'name': 'pTEF::KanMX::tTef', 'seq': 'TAGGTCTAGAGATCTGTTTAGCTTGCCTCGTCCCCGCCGGGTCACCCGGCCAGCGACATGGAGGCCCAGAATACCCTCCTTGACAGTCTTGACGTGCGCAGCTCAGGGGCATGATGTGACTGTCGCCCGTACATTTAGCCCATACATCCCCATGTATAATCATTTGCATCCATACATTTTGATGGCCGCACGGCGCGAAGCAAAAATTACGGCTCCTCGCTGCAGACCTGCGAGCAGGGAAACGCTCCCCTCACAGACGCGTTGAATTGTCCCCACGCCGCGCCCCTGTAGAGAAATATAAAAGGTTAGGATTTGCCACTGAGGTTCTTCTTTCATATACTTCCTTTTAAAATCTTGCTAGGATACAGTTCTCACATCACATCCGAACATAAACAACCATGGGTAAGGAAAAGACTCACGTTTCGAGGCCGCGATTAAATTCCAACATGGATGCTGATTTATATGGGTATAAATGGGCTCGCGATAATGTCGGGCAATCAGGTGCGACAATCTATCGATTGTATGGGAAGCCCGATGCGCCAGAGTTGTTTCTGAAACATGGCAAAGGTAGCGTTGCCAATGATGTTACAGATGAGATGGTCAGACTAAACTGGCTGACGGAATTTATGCCTCTTCCGACCATCAAGCATTTTATCCGTACTCCTGATGATGCATGGTTACTCACCACTGCGATCCCCGGCAAAACAGCATTCCAGGTATTAGAAGAATATCCTGATTCAGGTGAAAATATTGTTGATGCGCTGGCAGTGTTCCTGCGCCGGTTGCATTCGATTCCTGTTTGTAATTGTCCTTTTAACAGCGATCGCGTATTTCGTCTCGCTCAGGCGCAATCACGAATGAATAACGGTTTGGTTGATGCGAGTGATTTTGATGACGAGCGTAATGGCTGGCCTGTTGAACAAGTCTGGAAAGAAATGCATAAGCTTTTGCCATTCTCACCGGATTCAGTCGTCACTCATGGTGATTTCTCACTTGATAACCTTATTTTTGACGAGGGGAAATTAATAGGTTGTATTGATGTTGGACGAGTCGGAATCGCAGACCGATACCAGGATCTTGCCATCCTATGGAACTGCCTCGGTGAGTTTTCTCCTTCATTACAGAAACGGCTTTTTCAAAAATATGGTATTGATAATCCTGATATGAATAAATTGCAGTTTCATTTGATGCTCGATGAGTTTTTCTAATCAGTACTGACAATAAAAAGATTCTTGTTTTCAAGAACTTGTCATTTGTATAGTTTTTTTATATTGTAGTTGTTCTATTTTAATCAAATGTTAGCGTGATTTATATTTTTTTTCGCCTCGACATCATCTGCCCAGATGCGAAGTTAAGTGCGCAGAAAGTAATATCATGCGTCAATCGTATGTGAATGCTGGTCGCTATACTGCTGTCGATTCGATACTAACGCCGCCATCCAGTGTCGAAAACGAGCTCTCGAGAACCCTTAAT', 'need_reverse': 0, 'Primer_up_U': 'GV1', 'Primer_down_U': 'GV2', 'Primer_up': 'CGTGCGAuTAGGTCTAGAGATCTGTTTAGC', 'Primer_up_Tm': 50.26913247948829, 'Primer_down': 'CACGCGAuATTAAGGGTTCTCGAGAGC', 'Primer_down_Tm': 50.05341435932132}]\n"
     ]
    },
    {
     "data": {
      "text/html": [
       "<div>\n",
       "<style scoped>\n",
       "    .dataframe tbody tr th:only-of-type {\n",
       "        vertical-align: middle;\n",
       "    }\n",
       "\n",
       "    .dataframe tbody tr th {\n",
       "        vertical-align: top;\n",
       "    }\n",
       "\n",
       "    .dataframe thead th {\n",
       "        text-align: right;\n",
       "    }\n",
       "</style>\n",
       "<table border=\"1\" class=\"dataframe\">\n",
       "  <thead>\n",
       "    <tr style=\"text-align: right;\">\n",
       "      <th></th>\n",
       "      <th>Seq</th>\n",
       "      <th>Dir</th>\n",
       "      <th>Tm (c)</th>\n",
       "      <th>User</th>\n",
       "      <th>Template</th>\n",
       "      <th>Template_Len</th>\n",
       "      <th>benchling</th>\n",
       "    </tr>\n",
       "    <tr>\n",
       "      <th>Primer Name</th>\n",
       "      <th></th>\n",
       "      <th></th>\n",
       "      <th></th>\n",
       "      <th></th>\n",
       "      <th></th>\n",
       "      <th></th>\n",
       "      <th></th>\n",
       "    </tr>\n",
       "  </thead>\n",
       "  <tbody>\n",
       "    <tr>\n",
       "      <th>XX.0</th>\n",
       "      <td>CGTGCGAuTAGGTCTAGAGATCTGTTTAGC</td>\n",
       "      <td>fwd</td>\n",
       "      <td>50.27</td>\n",
       "      <td>GV1</td>\n",
       "      <td>pTEF::KanMX::tTef</td>\n",
       "      <td>1473</td>\n",
       "      <td>CGTGCGAtTAGGTCTAGAGATCTGTTTAGC</td>\n",
       "    </tr>\n",
       "    <tr>\n",
       "      <th>XX.0</th>\n",
       "      <td>CACGCGAuATTAAGGGTTCTCGAGAGC</td>\n",
       "      <td>rvs</td>\n",
       "      <td>50.05</td>\n",
       "      <td>GV2</td>\n",
       "      <td>pTEF::KanMX::tTef</td>\n",
       "      <td>1473</td>\n",
       "      <td>CACGCGAtATTAAGGGTTCTCGAGAGC</td>\n",
       "    </tr>\n",
       "  </tbody>\n",
       "</table>\n",
       "</div>"
      ],
      "text/plain": [
       "                                        Seq  Dir Tm (c) User  \\\n",
       "Primer Name                                                    \n",
       "XX.0         CGTGCGAuTAGGTCTAGAGATCTGTTTAGC  fwd  50.27  GV1   \n",
       "XX.0            CACGCGAuATTAAGGGTTCTCGAGAGC  rvs  50.05  GV2   \n",
       "\n",
       "                      Template Template_Len                       benchling  \n",
       "Primer Name                                                                  \n",
       "XX.0         pTEF::KanMX::tTef         1473  CGTGCGAtTAGGTCTAGAGATCTGTTTAGC  \n",
       "XX.0         pTEF::KanMX::tTef         1473     CACGCGAtATTAAGGGTTCTCGAGAGC  "
      ]
     },
     "execution_count": 60,
     "metadata": {},
     "output_type": "execute_result"
    }
   ],
   "source": [
    "\n",
    "\n",
    "KanMX={}\n",
    "KanMX['name']='pTEF::KanMX::tTef'\n",
    "KanMX['seq']='TAGGTCTAGAGATCTGTTTAGCttgcctcgtccccgccgggtcacccggccagcgacatggaggcccagaataccctccttgacagtcttgacgtgcgcagctcaggggcatgatgtgactgtcgcccgtacatttagcccatacatccccatgtataatcatttgcatccatacattttgatggccgcacggcgcgaagcaaaaattacggctcctcgctgcagacctgcgagcagggaaacgctcccctcacagacgcgttgaattgtccccacgccgcgcccctgtagagaaatataaaaggttaggatttgccactgaggttcttctttcatatacttccttttaaaatcttgctaggatacagttctcacatcacatccgaacataaacaaccatgggtaaggaaaagactcacgtttcgaggccgcgattaaattccaacatggatgctgatttatatgggtataaatgggctcgcgataatgtcgggcaatcaggtgcgacaatctatcgattgtatgggaagcccgatgcgccagagttgtttctgaaacatggcaaaggtagcgttgccaatgatgttacagatgagatggtcagactaaactggctgacggaatttatgcctcttccgaccatcaagcattttatccgtactcctgatgatgcatggttactcaccactgcgatccccggcaaaacagcattccaggtattagaagaatatcctgattcaggtgaaaatattgttgatgcgctggcagtgttcctgcgccggttgcattcgattcctgtttgtaattgtccttttaacagcgatcgcgtatttcgtctcgctcaggcgcaatcacgaatgaataacggtttggttgatgcgagtgattttgatgacgagcgtaatggctggcctgttgaacaagtctggaaagaaatgcataagcttttgccattctcaccggattcagtcgtcactcatggtgatttctcacttgataaccttatttttgacgaggggaaattaataggttgtattgatgttggacgagtcggaatcgcagaccgataccaggatcttgccatcctatggaactgcctcggtgagttttctccttcattacagaaacggctttttcaaaaatatggtattgataatcctgatatgaataaattgcagtttcatttgatgctcgatgagtttttctaatcagtactgacaataaaaagattcttgttttcaagaacttgtcatttgtatagtttttttatattgtagttgttctattttaatcaaatgttagcgtgatttatattttttttcgcctcgacatcatctgcccagatgcgaagttaagtgcgcagaaagtaatatcatgcgtcaatcgtatgtgaatgctggtcgctatactgctgtcgattcgatactaacgccgccatccagtgtcgaaaacgaGCTCTCGAGAACCCTTAAT'\n",
    "KanMX['need_reverse']=0 #if parts needs to be reversed:1 else:0\n",
    "KanMX['Primer_up_U']='GV1' #user site up\n",
    "KanMX['Primer_down_U']='GV2'#user site down\n",
    "KanMX['Primer_up']=''\n",
    "KanMX['Primer_up_Tm']=''\n",
    "KanMX['Primer_down']=''\n",
    "KanMX['Primer_down_Tm']=''\n",
    "\n",
    "#Design:\n",
    "importlib.reload(dt)\n",
    "Parts = dt.User_Assebmly([KanMX], 50, User_Overhangs)\n",
    "print(Parts)\n",
    "\n",
    "\n",
    "\n",
    "\n",
    "primer_name=[]\n",
    "primer_dir=[]\n",
    "primer_tm=[]\n",
    "primer_OH=[]\n",
    "primer_template_name=[]\n",
    "primer_seq=[]\n",
    "primer_template_len=[]\n",
    "benchling=[]\n",
    "i=1\n",
    "for part in Parts:\n",
    "    #here we adapt the order with which the overhang names are appended to match the Overhang names to the inversion of sequences\n",
    "    if part['need_reverse']==0:\n",
    "        primer_OH.append(part['Primer_up' +'_U'])\n",
    "        primer_OH.append(part['Primer_down'+'_U'])\n",
    "    elif part['need_reverse']==1:\n",
    "        primer_OH.append(part['Primer_down'+'_U'])\n",
    "        primer_OH.append(part['Primer_up' +'_U'])\n",
    "        \n",
    "    for primer in ['Primer_up', 'Primer_down']:\n",
    "        primer_name.append('XX.0')#.zfill fills primer number wiht leading 0\n",
    "        primer_seq.append(part[primer])\n",
    "        primer_tm.append(round(part[primer +'_Tm'],2))\n",
    "        primer_template_len.append(len(part['seq']))\n",
    "        primer_template_name.append(part['name'])\n",
    "        #primer_OH.append(part[primer +'_U'])\n",
    "        primer_dir.append('fwd'  if primer[7:] == 'up' else 'rvs' )\n",
    "        benchling.append(part[primer].replace('u','t'))\n",
    "        i+=1\n",
    "## need to fix the current overhang name bug (if seq needs inversion the OH name doenst match)\n",
    "        \n",
    "\n",
    "\n",
    "df = np.stack([primer_name, primer_seq,primer_dir, primer_tm,primer_OH, primer_template_name, primer_template_len, benchling] ).T\n",
    "Table=pd.DataFrame(df , columns=['Primer Name', 'Seq','Dir', 'Tm (c)','User', 'Template', 'Template_Len','benchling'])\n",
    "\n",
    "Table.set_index('Primer Name')"
   ]
  }
 ],
 "metadata": {
  "kernelspec": {
   "display_name": "DNAEnv",
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