Question: Quality Control Software For Targeted Sequencing?
3
gravatar for Luca Beltrame
6.7 years ago by
Luca Beltrame220
IRCCS Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy
Luca Beltrame220 wrote:

I'm mainly doing (amplicon-based) resequencing over small (50-100) number of genes with the Illumina technology. Aside from the normal metrics for quality control produced by Qualimap and FastQC, I'm wondering if there's something out there (aside TEQC, see below for while) that targets specifically targetd resequencing experiments.

I'm mostly interested in assessing coverage uniformity across a number of samples sequenced for my targets. I know about TEQC but it has a number of problems, in particular related to reading in paired reads (all of my experiments are paired-end), so I'm looking at alternatives.

Is there any software for QC that can be used for these types of experiments?

software ngs analysis sequencing • 2.5k views
ADD COMMENTlink modified 6.7 years ago by brentp23k • written 6.7 years ago by Luca Beltrame220
2
gravatar for brentp
6.7 years ago by
brentp23k
Salt Lake City, UT
brentp23k wrote:

picard's CollectTargetedPcrMetrics (http://picard.sourceforge.net/command-line-overview.shtml#CollectTargetedPcrMetrics) is a simple place to start.

It will tell you what % of your target bases are covered at 2X, 10X, 20X, 30X. What percent of your target bases are not covered and a few other useful metrics.

ADD COMMENTlink written 6.7 years ago by brentp23k

Looks interesting. Now all that's "left" is to find a way to convert Illumina's manifest format to Picard interval....

ADD REPLYlink written 6.7 years ago by Luca Beltrame220

to create that file, just do:

samtools view -H your.bam > intervals.list
awk 'BEGIN{FS=OFS="\t"}{ print $0"\t+\t"$1":"$2"-"$3 }' illumina.bed >> intervals.list
ADD REPLYlink written 6.7 years ago by brentp23k

Also in the latest versions of Picard there is a command that helps one to make such intervals from Bed files. They are: picard BedToIntervalList. This command requires a sequence dictionary that can be made using: picard CreateSequenceDictionary.

ADD REPLYlink written 19 months ago by Dataman330

The link above is broken. Here is a currently unbroken link to CollectTargetedPcrMetrics: https://software.broadinstitute.org/gatk/documentation/tooldocs/4.0.5.0/picard_analysis_directed_CollectTargetedPcrMetrics.php

ADD REPLYlink written 19 months ago by Dataman330
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