If ungapped alignment is softclipped, ksw_extend soft-clipped ends

It is possible that gapped alignment gives a better score than soft-clipping
based on ungapped alignment.

Closes #357

CHANGES.md

@@ -21,6 +21,9 @@
 * Include [ZStr](https://github.com/mateidavid/zstr/) in our own repository
   instead of downloading it at build time. This should make it possible to
   build strobealign without internet access.
+* #357: Fix some suboptimal alignment ends. Sometimes an end was soft-clipped
+  although a better alignment with an insertion or deletion existed that
+  extends to the end of the read.
 
 ## v0.12.0 (2023-11-23)
 

src/aligner.hpp

@@ -26,6 +26,7 @@ struct AlignmentInfo {
     int sw_score{0};
 
     int ref_span() const { return ref_end - ref_start; }
+    int query_span() const { return query_end - query_start; }
 };
 
 std::ostream& operator<<(std::ostream& os, const AlignmentInfo& info);

src/aln.cpp

@@ -222,11 +222,48 @@ inline Alignment extend_seed(
     int result_ref_start;
     bool gapped = true;
     if (projected_ref_end - projected_ref_start == query.size() && consistent_nam) {
-        std::string_view ref_segm_ham = ref.substr(projected_ref_start, query.size());
-        auto hamming_dist = hamming_distance(query, ref_segm_ham);
+        const std::string_view projected_ref = ref.substr(projected_ref_start, query.size());
+        info = hamming_align(query, projected_ref, aligner.parameters.match, aligner.parameters.mismatch, aligner.parameters.end_bonus);
+
+        if (info.edit_distance + (query.length() - info.query_span()) < 0.05 * query.length()) {
+            if (info.query_end < query.length()) {
+                // Right end is soft clipped, do gapped alignment on it
+                const std::string query_right = query.substr(info.query_end);
+                const int ext_right = std::min(ref.size() - projected_ref_end, size_t(50));
+                const std::string_view ref_right = ref.substr(projected_ref_start + info.ref_end, ext_right);
+                auto right = aligner.ksw_extend(query_right, ref_right, false);
+                info.query_end += right.query_end;
+                info.ref_end += right.ref_end;
+                info.edit_distance += right.edit_distance;
+                info.sw_score += right.sw_score;
+                assert(!info.cigar.empty());
+                info.cigar.pop_oplen();
+                info.cigar += right.cigar;
+            }
+
+            if (info.query_start > 0) {
+                // Left end is soft clipped, do gapped alignment on it
+                std::string query_left = query.substr(0, info.query_start);
+                const int ext_left = std::min(50, projected_ref_start);
+                const int ref_start = projected_ref_start - ext_left;
+                std::string ref_left{ref.substr(ref_start, ext_left + info.ref_start)};
+                std::reverse(query_left.begin(), query_left.end());
+                std::reverse(ref_left.begin(), ref_left.end());
+                auto left = aligner.ksw_extend(query_left, ref_left, true);
+                info.query_start -= left.query_end;
+                info.ref_start -= left.ref_end;
+                info.edit_distance += left.edit_distance;
+                info.sw_score += left.sw_score;
+
+                // TODO this just removes the soft-clipping from the beginning,
+                // a bit too complicated
+                info.cigar.reverse();
+                info.cigar.pop_oplen();
+                info.cigar += left.cigar;
+                info.cigar.reverse();
+            }
 
-        if (hamming_dist >= 0 && (((float) hamming_dist / query.size()) < 0.05) ) { //Hamming distance worked fine, no need to ksw align
-            info = hamming_align(query, ref_segm_ham, aligner.parameters.match, aligner.parameters.mismatch, aligner.parameters.end_bonus);
+            assert(info.cigar.derived_sequence_length() == query.length());
             result_ref_start = projected_ref_start + info.ref_start;
             gapped = false;
         }

src/cigar.hpp

@@ -93,6 +93,10 @@ class Cigar {
         std::reverse(m_ops.begin(), m_ops.end());
     }
 
+    void pop_oplen() {
+        m_ops.pop_back();
+    }
+
     /* Return a new Cigar that uses M instead of =/X */
     Cigar to_m() const;
 

tests/phix.1.fastq

@@ -31,9 +31,9 @@ NTCATTTGGCGAGAAAGCTCAGTCTCAGGAGGAAGCGGAGCAGGCCAAATGTTTTTGAGATGGCAGCAACGGAAACCATA
 +
 #88ABCFG<EC:CCCF<,6CFGE<;EFEGFE@;F,CFB7F7CF,6BCCCFFCFEE@CGCGGDCCECEFGC@@FGEEGGFGFC=?F@FCC=EGGF?CFFGAFFFFG<?FGF<EDFGC<FEG7CE=ECGGGG8EGFEFGGF+>=>+@+3,3CFB<CFGCF,EFGCGBFGGG9FGCFD:,@=@FFGGFF9CFGGGGGGGG6@CGCGGGGGGGGC7F:7CDB<CEGCGGGG46DC4CD37BCAFGFGAFA77AFFA;FAFC;CA:464<9<?#################################
 @SRR1377138.9
-NTTTGTTAACGTATTTAGCCACATAGAAACCAACAGCCATATAACTGGTAGCTTTAAGCGGCTCACCTTTAGCATCAACAGGCCACAACCAACCAGAACGTGAAAAAGCGTCCTGCGTGTAGCGAACTGCGATGGGCATACTGTAACCATAAGGCCACGTATTTTGCAAGCTATTTAACTGGCGGCGATTGCGTACCCGACGACCAAAATTAGGGTCAACGCTACCTGTAGGAAGTGTCCGCATAAAGTGCACCGCATGGAAATGAAGACGGCCATTAGCTGTACCATACTCAGGCACACA
+NTTTGTTAACGTATTTAGCCACATAGAAACCAACAGCCATATAACTGGTAGCTTTAAGCGGCTCACCTTTAGCATCAACAGGCCACAACCAACCAGAACGTGAAAAAGCGTCCTGCGTGTAGCGAACTGCGATGGGCATACTGTAACCATAAGGCCACGTATTTTGCAAGCTATTTAACTGGCGGCGATTGCGTACCCGACGACCAAAATTAGGGTCAACGCTACCTGTAGGAAGTGTCCGCATAAAGTGCACCGCATGGAAATGAAGACGGCCATTAGCTGTACCATACTCAGGACACA
 +
-#8BCCGGGGGGGGGGGGGGGCEFFGFGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFFFGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFGGGGGGGGGGGGGGGGGGGGGGGEGGGGGGGGGGGGGGGGGGGGGGGEGGGGGGGGGGGGEDGGGGGGGGGGFGGGFGFGGFGFGFF=7??FFFEBBFBFBBFFFFFFF:-044;?FFFFFFFFABFB?FFF<?:<?FFB?
+#8BCCGGGGGGGGGGGGGGGCEFFGFGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFFFGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFGGGGGGGGGGGGGGGGGGGGGGGEGGGGGGGGGGGGGGGGGGGGGGGEGGGGGGGGGGGGEDGGGGGGGGGGFGGGFGFGGFGFGFF=7??FFFEBBFBFBBFFFFFFF:-044;?FFFFFFFFABFB?FFF<?:?FFB?
 @SRR1377138.10
 NAAATGATTGGCGTATCCAACCTGCAGAGTTTTATCGCTTCCATGACGCAGAAGTTAACACTTTCGGATATTTCTGATGAGTCGAAAAATTATCTTGATAAAGCAGGAATTACTACTGCTTGTTTACGAATTAAATCGAAGTGGACTGCTGGCGGAAAATGAGAAAATTCGACCTATCCTTGCGCAGCTCGAGAAGCTCTTACTTTGCGACCTTTCGCCATCAACTAACGATTCTGTCAAAAACTGACGCGTTGGATGAGGAGAAGTGGCTTAATATGCTTGGCACGTTCGTCAAGGACTG
 +
@@ -47,9 +47,9 @@ NAATCTGCTCAAATTTATGCGCGCTTCGATAAAAATGATTGGCGTATCCAACCTGCAGAGTTTTATCGCTTCCATGACGC
 +
 #8BCCGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFGGGDCGGGGGGGGGGFGGGGGGGFGFGGFGFGFFFFFFFFFFFAFFFFFFFFFFFA3.1.48?A<:?FFFFFFBDFFFF?1<BAFFF#
 @SRR1377138.13
-NTATGTGGCTAAATACGTTAACAAAAAGTCAGATATGGACCTTGCTGCTAAAGGTCTAGGAGCTAAAGAATGGAACAACTCACTAAAAACCAAGCTGTCGCTACTTCCCAAGAAGCTGTTCAGAATCAGAATGAGCCGCAACTTCGGGATGAAAATGCTCACAATGACAAATCTGTCCACGGAGTGCTTAATCCAACTTACCAAGCTGGGTTACGACGCGACGCCGTTCAACCAGATATTGAAGCAGAACGCAAAAAGAGAGATGAGATTGAGGCTGGGAAAAGTTACTGTAGCCGACGTT
+NTATGTGGCTAAATACGTTAACAAAAAGTCAGATATGGACCTTGCTGCTAAAGGTCTAGGAGCTAAAGAATGGAACAACTCACTAAAAACCAAGCTGTCGCTACTTCCCAAGAAGCTGTTCAGAATCAGAATGAGCCGCAACTTCGGGATGAAAATGCTCACAATGACAAATCTGTCCACGGAGTGCTTAATCCAACTTACCAAGCTGGGTTACGACGCGACGCCGTTCAACCAGATATTGAAGCAGAACGCAAAAAGAGAGATGAGATTGAGGCTGGGAAAAGTTACTGTAGCGACGTT
 +
-#8BCCGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFCFGGFGFGFFFFFFFFFFFDFFFFB:>FFFFF2.1.4::?FFFFF?BAAFFFFFFDFF>>9>9?
+#8BCCGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFCFGGFGFGFFFFFFFFFFFDFFFFB:>FFFFF2.1.4::?FFFFF?BAAFFFFFFDF>>9>9?
 @SRR1377138.14
 NTCTTGATAAAGCAGGAATTACTACTGCTTGTTTACGAATTAAATCGAAGTGGACTGCTGGCGGAAAATGAGAAAATTCGACCTATCCTTGCGCAGCTCGAGAAGCTCTTACTTTGCGACCTTTCGCCATCAACTAACGATTCTGTCAAAAACTGACGCGTTGGATGAGGAGAAGTGGCTTAATATGCTTGGCACGTTCGTCAAGGACTGGTTTAGATATGAGTCACATTTTGTTCATGGTAGAGATTCTCTTGTTGACATTTTAAAAGAGCGTGGATTACTATCTGAGTCCGATGCTGTT
 +

tests/phix.pe.sam

@@ -17,15 +17,15 @@ SRR1377138.7	83	NC_001422.1	256	60	300=1X	=	141	-416	GGCACGTTCGTCAAGGACTGGTTTAGA
 SRR1377138.7	163	NC_001422.1	141	60	1X299=1X	=	256	416	NTTCGACCTATCCTTGCGCAGCTCGAGAAGCTCTTACTTTGCGACCTTTCGCCATCAACTAACGATTCTGTCAAAAACTGACGCGTTGGATGAGGAGAAGTGGCTTAATATGCTTGGCACGTTCGTCAAGGACTGGTTTAGATATGAGTCACATTTTGTTCATGGTAGAGATTCTCTTGTTGACATTTTAAAAGAGCGTGGATTACTATCTGAGTCCGATGCTGTTCAACCACTAATAGGTAAGAAATCATGAGTCAAGTTACTGAACAATCCGTACGTTTCCAGACCGCTTTGGCCTCTT	#8A<BFCFEGFFGGF9C:CFCEGGCCGECFF@EEGGGGGGFGCEEGEFGEGGECFCFGFGG<<F@@EFFEFGGDCFGGCFGGC:CB:68AA,BBC=FDA8FAFG8=FFFFEFCF<A+A??=FGCCFEFG:?F=AFDGGGGGFCFFFFCFGGGGGFGGFGGFC,7DDFFFFGGGGGFFEGGGFGGGGGG@7;8=+=36=DEDCABFF>F6BFFCFFF4AA>A4A4>:E>7?-55>;(>?E38*878CB6@EC7C;6C>C<)6<C))))/;;7459,2;>FFF@E>2)(4179:BB?FB####	NM:i:2	AS:i:602	RG:Z:1
 SRR1377138.8	83	NC_001422.1	1254	60	124=1X132=1X42=1X	=	1179	-376	TCATGAAGGATGGTGTTAATGCCACTCCTCTCCCGACTGTTAACACTACTGGTTATATTGACCATGCCGCTTTTCTTGGCACGATTAACCCTGATACCAATAAAATCCCTAAGCATTTGTTTCATGGTTATTTGAATATCTATAACAACTATTTTAAAGCGCCGTGGATGCCTGACCGTACCGAGGCTAACCCTAATGAGCTTAATCAAGATGATGCTCGTTATGGTTTCCGTTGCTGCCATCTCAAAAACATTTGGCCTGCTCCGCTTCCTCCTGAGACTGAGCTTTCTCGCCAAATGAN	#################################?<9<464:AC;CFAF;AFFA77AFAGFGFACB73DC4CD64GGGGCGEC<BDC7:F7CGGGGGGGGCGC@6GGGGGGGGFC9FFGGFF@=@,:DFCGF9GGGFBGCGFE,FCGFC<BFC3,3+@+>=>+FGGFEFGE8GGGGCE=EC7GEF<CGFDE<FGF?<GFFFFAGFFC?FGGE=CCF@F?=CFGFGGEEGF@@CGFECECCDGGCGC@EEFCFFCCCB6,FC7F7BFC,F;@EFGEFE;<EGFC6,<FCCC:CE<GFCBA88#	NM:i:3	AS:i:592	RG:Z:1
 SRR1377138.8	163	NC_001422.1	1179	60	1X79=1X112=1X88=1X15=1X2=	=	1254	376	NTATTGACTCTACTGTAGACATTTTTACTTTTTATGTCCCTCATCGTCACGTTTATGGTGAACAGTGGATTAAGTTCATGCAGGATGGTGTTAATGCCACTCCTCTCCCGACTGTTAACACTACTGGTTATATTGACCATGCCGCTTTTCTTGGCACGATTAACCCTGATACCAATAAAATCCCTAAGCATTTTTTTCAGGGTTATTTGAATATCTATAACAACTATTTTAAAGCGCCGTGGATGCCTGACCGTACCGAGGCTAACCCTAATGAGCTTAATCCAGATGATGCTCGTTACGG	#8AAC,6C;EC@E,@FA,<CC<F,,CFFCGGEGAEECFGFFGG<E@@@@F:CCA<F<EF,@;;CC@@EC<FFGDFEF@<,,66,,B<,F<BC<5F@,,95BEFD,?9FEFEFGGCFE?BE=FC=?FFF:;AFCFEG;E=FGFFBE+CFEFEG9=B?FGGGGDD89=@DGFGGGD8DDGFFD?8A8,=+8=FFD+?DDFD+98F?F8C?+CF;3?===F7C7?@@)=A0@EF)*>EFF5>A@FFE=<)1*-..:@29@29@0:1;3=6@=(544?4)9;C>5))7).474CB)99?######	NM:i:5	AS:i:572	RG:Z:1
-SRR1377138.9	83	NC_001422.1	4731	60	300=1X	=	4650	-382	TGTGTGCCTGAGTATGGTACAGCTAATGGCCGTCTTCATTTCCATGCGGTGCACTTTATGCGGACACTTCCTACAGGTAGCGTTGACCCTAATTTTGGTCGTCGGGTACGCAATCGCCGCCAGTTAAATAGCTTGCAAAATACGTGGCCTTATGGTTACAGTATGCCCATCGCAGTTCGCTACACGCAGGACGCTTTTTCACGTTCTGGTTGGTTGTGGCCTGTTGATGCTAAAGGTGAGCCGCTTAAAGCTACCAGTTATATGGCTGTTGGTTTCTATGTGGCTAAATACGTTAACAAAN	?BFF?<:?<FFF?BFBAFFFFFFFF?;440-:FFFFFFFBBFBFBBEFFF??7=FFGFGFGGFGFGGGFGGGGGGGGGGDEGGGGGGGGGGGGEGGGGGGGGGGGGGGGGGGGGGGGEGGGGGGGGGGGGGGGGGGGGGGGFGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFFFGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFGFFECGGGGGGGGGGGGGGGCCB8#	NM:i:1	AS:i:612	RG:Z:1
+SRR1377138.9	83	NC_001422.1	4731	60	5=1D294=1X	=	4650	-382	TGTGTCCTGAGTATGGTACAGCTAATGGCCGTCTTCATTTCCATGCGGTGCACTTTATGCGGACACTTCCTACAGGTAGCGTTGACCCTAATTTTGGTCGTCGGGTACGCAATCGCCGCCAGTTAAATAGCTTGCAAAATACGTGGCCTTATGGTTACAGTATGCCCATCGCAGTTCGCTACACGCAGGACGCTTTTTCACGTTCTGGTTGGTTGTGGCCTGTTGATGCTAAAGGTGAGCCGCTTAAAGCTACCAGTTATATGGCTGTTGGTTTCTATGTGGCTAAATACGTTAACAAAN	?BFF?:?<FFF?BFBAFFFFFFFF?;440-:FFFFFFFBBFBFBBEFFF??7=FFGFGFGGFGFGGGFGGGGGGGGGGDEGGGGGGGGGGGGEGGGGGGGGGGGGGGGGGGGGGGGEGGGGGGGGGGGGGGGGGGGGGGGFGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFFFGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFGFFECGGGGGGGGGGGGGGGCCB8#	NM:i:2	AS:i:597	RG:Z:1
 SRR1377138.9	163	NC_001422.1	4650	60	1X253=1X45=1X	=	4731	382	NTTCGTGATATTGGTCGTATGGTTCTTGCTGCCGAGGGTCGCAAGGCTAATGATTCACACGCCGACTGCTATCAGTATTTTTGTGTGCCTGAGTATGGTACAGCTAATGGCCGTCTTCATTTCCATGCGGTGCACTTTATGCGGACACTTCCTACAGGTAGCGTTGACCCTAATTTTGGTCGTCGGGTACGCAATCGCCGCCAGTTAAATAGCTTGCAAAATACGTGGCCTTATGGTTACAGTATGCCCATCGCCGTTCGCTACACGCAGGACGCTTTTTCACGTTCTGGTTGGTTGTGGG	#8BCCGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFGGGGGGGGGGGGGGDGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG9EFGEGEGGGGGGGGGGFFGGGGGGGGGGGGGGGGGGGAEGGGGGGGGGGGGGGGGGFGGFFG;BAFFFAFFFFFFF>FCF@FFF?EFFF@EFEFFFFEEEB?FD<<)8083:<F((4:??01,72??##################################	NM:i:3	AS:i:592	RG:Z:1
 SRR1377138.10	99	NC_001422.1	1	60	26S275=	=	58	358	NAAATGATTGGCGTATCCAACCTGCAGAGTTTTATCGCTTCCATGACGCAGAAGTTAACACTTTCGGATATTTCTGATGAGTCGAAAAATTATCTTGATAAAGCAGGAATTACTACTGCTTGTTTACGAATTAAATCGAAGTGGACTGCTGGCGGAAAATGAGAAAATTCGACCTATCCTTGCGCAGCTCGAGAAGCTCTTACTTTGCGACCTTTCGCCATCAACTAACGATTCTGTCAAAAACTGACGCGTTGGATGAGGAGAAGTGGCTTAATATGCTTGGCACGTTCGTCAAGGACTG	#8BCCGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGDGGGGGGGGGGGGGGGGGGGGFBFFGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFGFGGFGFGFFFFFFFFFFFFFFFFFF?BFFFF2.8:<BFFFFFFFFFFBFFFBF?BBF???F?B	NM:i:0	AS:i:560	RG:Z:1
 SRR1377138.10	147	NC_001422.1	58	60	300=1X	=	1	-358	GAAAAATTATCTTGATAAAGCAGGAATTACTACTGCTTGTTTACGAATTAAATCGAAGTGGACTGCTGGCGGAAAATGAGAAAATTCGACCTATCCTTGCGCAGCTCGAGAAGCTCTTACTTTGCGACCTTTCGCCATCAACTAACGATTCTGTCAAAAACTGACGCGTTGGATGAGGAGAAGTGGCTTAATATGCTTGGCACGTTCGTCAAGGACTGGTTTAGATATGAGTCACATTTTGTTCATGGTAGAGATTCTCTTGTTGACATTTTAAAAGAGCGTGGATTACTATCTGAGTCCN	###FBDA<5)2<BAAFBA>?>FFDFFECC49FFD?3<@FFFFEDEC?3E?C?0<FFFFFFFEFFDFBFFFFFFFFE6E;FFFFFF>EFFFFFFF>@5FECFFGGGFGD:FGGCDGFGGGGGG>GGGGGGGDGGGGGGGGGGGGGGGGFGGGGGGGGGGGGGGGEGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGCCB8#	NM:i:1	AS:i:612	RG:Z:1
 SRR1377138.11	99	NC_001422.1	5117	60	1X268=32S	=	5220	270	NAAGCTGTTCAGAATCAGAATGAGCCGCAACTTCGGGATGAAAATGCTCACAATGACAAATCTGTCCACGGAGTGCTTAATCCAACTTACCAAGCTGGGTTACGACGCGACGCCGTTCAACCAGATATTGAAGCAGAACGCAAAAAGAGAGATGAGATTGAGGCTGGGAAAAGTTACTGTAGCCGACGTTTTGGCGGCGCAACCTGTGACGACAAATCTGCTCAAATTTATGCGCGCTTCGATAAAAATGATTGGCGTATCCAACCTGCGGAGTTTTATCGCTTCCATGACGCAGAAGTTA	#8ACCGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFFGGGGGGGGGGGGGGGGGGGGGGGGGGFGGGGGGGGGGGGGGGGGGGGGGGGFGGGGGGGGGDGGGGF8FFGGGGGGGGGGGGGGGGGGGFGGGGGGGGGGGGGGGGGFGGGCFGGGGGGGGGGEE5EGGGGGGGGGFGGGGFDGEGDGGDGGGE7FFGGG>*?*:<>9BDECDFGF<*9@F?*7B?;F;F;B:AFFFFFF7*(03:0:AAAFFF:18?BA:<B##########	NM:i:1	AS:i:538	RG:Z:1
 SRR1377138.11	147	NC_001422.1	5220	60	167=134S	=	5117	-270	GACGCGACGCCGTTCAACCAGATATTGAAGCAGAACGCAAAAAGAGAGATGAGATTGAGGCTGGGAAAAGTTACTGTAGCCGACGTTTTGGCGGCGCAACCTGTGACGACAAATCTGCTCAAATTTATGCGCGCTTCGATAAAAATGATTGGCGTATCCAACCTGCAGAGTTTTATCGCTTCCATGACGCAGAAGTTAACACTTTCGGATATTTCTGATGAGTCGAAAAATTATCTTGATAAAGCAGGAATTACTACTGCTTGTTTACGAATTAAATCGAAGTGGACTGCTGGCGGAAAAN	########BFFBA>82><9B>4<2BAA2A<<:?:4,.(B?C?.37;)EE<;)C?6(((;FFFFFEE?7)?9/);FDFB>A=FDAFFFFFFFDEDDCCFAGFFGGGGGGF:GGGFGGGGGFGGGGFGCGGEECDEFCCGGFGGGGFGGGGGGGGGGGGGGGGGFGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFGGGFFGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGCCA8#	NM:i:0	AS:i:344	RG:Z:1
 SRR1377138.12	99	NC_001422.1	1	60	57S238=1X5=	=	28	328	NAATCTGCTCAAATTTATGCGCGCTTCGATAAAAATGATTGGCGTATCCAACCTGCAGAGTTTTATCGCTTCCATGACGCAGAAGTTAACACTTTCGGATATTTCTGATGAGTCGAAAAATTATCTTGATAAAGCAGGAATTACTACTGCTTGTTTACGAATTAAATCGAAGTGGACTGCTGGCGGAAAATGAGAAAATTCGACCTATCCTTGCGCAGCTCGAGAAGCTCTTACTTTGCGACCTTTCGCCATCAACTAACGATTCTGTCAAAAACTGACGCGTTGGATGAGGAGATGTGGC	#8BCCGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFGGGDCGGGGGGGGGGFGGGGGGGFGFGGFGFGFFFFFFFFFFFAFFFFFFFFFFFA3.1.48?A<:?FFFFFFBDFFFF?1<BAFFF#	NM:i:1	AS:i:488	RG:Z:1
 SRR1377138.12	147	NC_001422.1	28	60	211=1X88=1X	=	1	-328	GTTAACACTTTCGGATATTTCTGATGAGTCGAAAAATTATCTTGATAAAGCAGGAATTACTACTGCTTGTTTACGAATTAAATCGAAGTGGACTGCTGGCGGAAAATGAGAAAATTCGACCTATCCTTGCGCAGCTCGAGAAGCTCTTACTTTGCGACCTTTCGCCATCAACTAACGATTCTGTCAAAAACTGACGCGTTGGATGAGGAGATGTGGCTTAATATGCTTGGCACGTTCGTCAAGGACTGGTTTAGATATGAGTCACATTTTGTTCATGGTAGAGATTCTCTTGTTGACATTN	#FFFA<600B??A<4FA?BF?AFB?21DD9FFFFFECEFE<CFECEFFFFFFFFFEE?FFFFFFBBFFC9BFFFFFFFCFEFDBFFFFFFFFFFFFFFFGFFGGGGGF>GGGGGGGGGGGGGGGGGGGGGGGGGGGGGFGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFFGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGCCB8#	NM:i:2	AS:i:602	RG:Z:1
-SRR1377138.13	99	NC_001422.1	5006	60	1X300=	=	5049	344	NTATGTGGCTAAATACGTTAACAAAAAGTCAGATATGGACCTTGCTGCTAAAGGTCTAGGAGCTAAAGAATGGAACAACTCACTAAAAACCAAGCTGTCGCTACTTCCCAAGAAGCTGTTCAGAATCAGAATGAGCCGCAACTTCGGGATGAAAATGCTCACAATGACAAATCTGTCCACGGAGTGCTTAATCCAACTTACCAAGCTGGGTTACGACGCGACGCCGTTCAACCAGATATTGAAGCAGAACGCAAAAAGAGAGATGAGATTGAGGCTGGGAAAAGTTACTGTAGCCGACGTT	#8BCCGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFCFGGFGFGFFFFFFFFFFFDFFFFB:>FFFFF2.1.4::?FFFFF?BAAFFFFFFDFF>>9>9?	NM:i:1	AS:i:612	RG:Z:1
+SRR1377138.13	99	NC_001422.1	5006	60	1X293=1D6=	=	5049	344	NTATGTGGCTAAATACGTTAACAAAAAGTCAGATATGGACCTTGCTGCTAAAGGTCTAGGAGCTAAAGAATGGAACAACTCACTAAAAACCAAGCTGTCGCTACTTCCCAAGAAGCTGTTCAGAATCAGAATGAGCCGCAACTTCGGGATGAAAATGCTCACAATGACAAATCTGTCCACGGAGTGCTTAATCCAACTTACCAAGCTGGGTTACGACGCGACGCCGTTCAACCAGATATTGAAGCAGAACGCAAAAAGAGAGATGAGATTGAGGCTGGGAAAAGTTACTGTAGCGACGTT	#8BCCGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGFCFGGFGFGFFFFFFFFFFFDFFFFB:>FFFFF2.1.4::?FFFFF?BAAFFFFFFDF>>9>9?	NM:i:2	AS:i:597	RG:Z:1
 SRR1377138.13	147	NC_001422.1	5049	60	300=1X	=	5006	-344	GCTGCTAAAGGTCTAGGAGCTAAAGAATGGAACAACTCACTAAAAACCAAGCTGTCGCTACTTCCCAAGAAGCTGTTCAGAATCAGAATGAGCCGCAACTTCGGGATGAAAATGCTCACAATGACAAATCTGTCCACGGAGTGCTTAATCCAACTTACCAAGCTGGGTTACGACGCGACGCCGTTCAACCAGATATTGAAGCAGAACGCAAAAAGAGAGATGAGATTGAGGCTGGGAAAAGTTACTGTAGCCGACGTTTTGGCGGCGCAACCTGTGACGACAAATCTGCTCAAATTTATGN	#FFFFBFFFBD>A<::64>AAFFFB><<A>?FFBABBAFFFFFFBAFFFFFFFFFFFFFE;@F@FFFFFFFD@FE?FFFFFFFFFFFFFEBFFFFFFFFGFFGGGDGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGDGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGCGGCGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGCCC8#	NM:i:1	AS:i:612	RG:Z:1
 SRR1377138.14	99	NC_001422.1	66	60	1X300=	=	156	391	NTCTTGATAAAGCAGGAATTACTACTGCTTGTTTACGAATTAAATCGAAGTGGACTGCTGGCGGAAAATGAGAAAATTCGACCTATCCTTGCGCAGCTCGAGAAGCTCTTACTTTGCGACCTTTCGCCATCAACTAACGATTCTGTCAAAAACTGACGCGTTGGATGAGGAGAAGTGGCTTAATATGCTTGGCACGTTCGTCAAGGACTGGTTTAGATATGAGTCACATTTTGTTCATGGTAGAGATTCTCTTGTTGACATTTTAAAAGAGCGTGGATTACTATCTGAGTCCGATGCTGTT	#8@CCGGGGGGFFGGGGGGGGGGGGGFGGGGGGGGGGGGGFGGEEFGGGGGGGGGGGGGFGDCFEFGGGGGFGGGGGGGGGGGGGGGGGGGG7FGGGGGGGGGGGGGGGGGGGDFGGGGGGGFGGEGGGGGFGGGGGGGGGGGGGGGGGFGGGGGGGGGGDGGGGGGGGDGGGGGGGFEGGGGGGGFGGFFGFGGGGGGGGGFGGGFG7FGGFGDGGGGGFFCGGGGGGGGGGGGGFGFGGC>FGCF=A@FFFFFFFFFFEFFFFFFE<=/04(34:5<BFFA<AFFAAABFF>01:AB>4	NM:i:1	AS:i:612	RG:Z:1
 SRR1377138.14	147	NC_001422.1	156	60	300=1X	=	66	-391	GCGCAGCTCGAGAAGCTCTTACTTTGCGACCTTTCGCCATCAACTAACGATTCTGTCAAAAACTGACGCGTTGGATGAGGAGAAGTGGCTTAATATGCTTGGCACGTTCGTCAAGGACTGGTTTAGATATGAGTCACATTTTGTTCATGGTAGAGATTCTCTTGTTGACATTTTAAAAGAGCGTGGATTACTATCTGAGTCCGATGCTGTTCAACCACTAATAGGTAAGAAATCATGAGTCAAGTTACTGAACAATCCGTACGTTTCCAGACCGCTTTGGCCTCTATTAAGCTCATTCAGN	############?>BFAAA45FFFFB:7/(9>9;B@=63?FFED>52FE?>5*>852BCEEFFAA@A>8@8C>=7EFCFFEAFFFF:FAFFFFFFFFD?9*DDGGGGGDGGGGGGFGFGGFFCFFGDGFGGGGGFGFEF?GGFCGGGGGGGGGGGGFEEGFFGGFE9GGGFFGFAGGGGGGGGFAGGGFGFFFGGGGGGGEFAFFCFGGGF@GF@AE@FFCEF<FGFGFDGGGGEEFAFGAGGFFFFCGFCFGGFDF@FEEGGE@C,GF@CCCF@CGGGFEEC,GGFF@FFEF<FF@@A8#	NM:i:1	AS:i:612	RG:Z:1