calculate haloplex coverage

Question

[jonessara770]

Hi,

How do you calculate the coverage of target genes with Haloplex data? I am trying to use GATK but it gives 0 for TP53. However, when I check the coverage in IGV the gene is well covered...

any suggestion?

Thanks Sara

[Pierre Lindenbaum]

a few questions:

1) what is the output of

samtools view -c your.bam input "TP53chrom:TP53star-TP53end" ?

2) what was your gatk command line ? 3) did you remove the PCR duplicate like picard "Markduplicate" ?

[jonessara770]

Thanks 1) this command samtools view -c your.bam input "TP53chrom:TP53star-TP53end" ? gives me

22372

2) java -jar GenomeAnalysisTK.jar -T DepthOfCoverage -R Homo_sapiens.GRCh37.fa -I mybam.matefixed.sorted.bam -L gene.bed -o myout.genelist_coverage --minMappingQuality 15 --minBaseQuality 10 --logging_level INFO --summaryCoverageThreshold 5 --summaryCoverageThreshold 7 --summaryCoverageThreshold 10 --summaryCoverageThreshold 15 --summaryCoverageThreshold 20 --summaryCoverageThreshold 30 --summaryCoverageThreshold 50

3) I did not remove duplicate coz it is a Haloplex and it is recommended by the company to not remove duplicates

[Pierre Lindenbaum]

are you sure about gene.bed (same build ?) try

  samtools view -c -L gene.bed mybam.matefixed.sorted.bam

[jonessara770]

Here is the output : 22372

Do you recommend any other tools to get the coverage? thanks

[Pierre Lindenbaum]

no idea. evrythin looks fine... all the reads have a bad quality ? are poorly mapped (orphan) ? ...

[jonessara770]

How can I check these?

Answer 1

[trausch]

We have used Alfred to calculate Haloplex on-target rate and target coverage distribution.

alfred qc -r <ref.fa> -b <targets.bed.gz> <align.bam>