No genotype calls for well-covered locus

[jlac]

Hi,

I am trying to genotype TRs at the FXN locus and am getting strange results for some samples where the genotypes are not called at all. I know in a previous thread this was attributed to inadequate coverage, but in the attached plot you can see good coverage across several samples, but CloneA receives a call and CloneB does not.

I am wondering if it would be possible to improve the TRGT outputs to give some information about why a particular location wasn't genotyped? Or alternatively, if all sites can be genotyped but sites that don't meet minimum thresholds are not marked as PASS. Without these types of info it's almost impossible to understand why a particular sample can be genotyped while another cannot.

Thank you!

[pbsena]

Hello,

Thanks for bringing up the question! The most recent version (0.7.0) of trgt contains some advanced parameters that can be tweaked to investigate the lack of coverage. If you run it with verbose option (-v), you may see something like this:

2024-01-19 19:08:49 [DEBUG] - FXN: Collected 506 reads
2024-01-19 19:08:49 [DEBUG] - FXN: 506 reads left after clipping
2024-01-19 19:09:14 [DEBUG] - FXN: Found 475 spanning reads
2024-01-19 19:09:14 [DEBUG] - FXN: 474 spanning reads had sufficiently long flanks
2024-01-19 19:09:14 [DEBUG] - FXN: downsampled to 474 reads

If you are losing reads at the "long flanks" step, I'd suggest trying to tweak the --min-flank-id-frac value
if you see a warning such as this:

2024-01-19 19:13:23 [WARN] - Quality filtered 321 out of 332 reads

I'd suggest rerunning with --min-read-quality=-1

Hopefully those can shed light on the lack of coverage, but otherwise I'd love to reiterate either based on the verbose outputs you get or, if needed, looking at the BAM files that are causing issues.

[jlac]

Hi,

Thank you for the reply! I ran it with the --verbose flag and am not getting the additional DEBUG details you outlined above.

[pbsena]

@jlac can you try instead adding three -v commands (i.e.,: trgt -v -v -v) instead of just --verbose? This should print the debug info.

[jlac]

That worked! Thank you!