Quality check of NGS data

Question

[stanedav]

Hello, fellow bioinformaticians,

I have a question about a quality check of NGS data. I was asked to perform a quality check on NGS data in our lab. We are outsourcing sequencing, so we receiving FASTQ from DNA sample - mostly WES.

Then with WES data, I am performing virtual panel analysis (alignment and variant calling with custom bed file), then open WES (with prepared bed file).

We need to report the quality check of the data (FASTQ, BAM?) and in the case of virtual panel summary about coverage of genes in the virtual panel. Ideally, the report should be one sheet of paper.

Which tools would you use? I am using now bamstats for coverage of genes and exons.

I also tried MultiQC and Alfred tool, are these two ok?

Answer 1

[Bastien Hervé]

Hello,

There are various tools to have a look at sequencing quality, you can try MultiQC or fastp for example to have a look at reads quality, reads length, contamination, over expressed sequences...

For alignment quality check, you can try to have a look at alignment falling outside your gene panel. In order to do so you can take a look at bedtools, something like :

bedtools intersect -v -abam the.bam -b probes.bed

Finding The Reads Mapped Outside The Target Region

Answer 2

[iraun]

Hey!

• For fastq quality control I would recommend the tradicional well known FASTQC tool
• For BAM, you can use as you mentioned, bamstat, or another alternative can be Qualimap

MultiQC can take the output of all of them and create a comprehensive report. Another tool that I personally like and use often is mosdepth.

Hope it helps!