Question: Questions regarding whole exome sequencing
0
gravatar for nickhir
5 weeks ago by
nickhir10
german research cancer center (DKFZ)
nickhir10 wrote:

Hey everybody, I am currently informing myself about Whole exome sequencing (WES) and there are a couple of things that still confuse me:

1) I read that WES "does not permit deep sequencing". Can somebody elaborate why this is the case?

2) I checked this website to get an idea about coverage for WES and I did not understand, why it is necessary to have a much higher coverage for detecting SNV when using WES compared to whole genome sequencing (e.g. 100x vs 15x).

If anyone could answer these questions for me, I'd be very thankful!!

Thanks!

sequencing wes • 90 views
ADD COMMENTlink modified 5 weeks ago by Kevin Blighe61k • written 5 weeks ago by nickhir10
0
gravatar for Kevin Blighe
5 weeks ago by
Kevin Blighe61k
Kevin Blighe61k wrote:

1) I read that WES "does not permit deep sequencing". Can somebody elaborate why this is the case?

One can have relatively high depth of coverage even with whole genome sequencing. However, factors that limit depth of coverage include:

  • input DNA amount and concentration
  • input DNA quality
  • sequencer used and flow-cell capacity
  • number of sequencing cycles used
  • library prep used
  • reagent availability
  • et cetera

Coverage over certain regions of the genome will also always be limited due to GC content, sequence similarity, and other factors.

2) I checked this website to get an idea about coverage for WES and I did not understand, why it is necessary to have a much higher coverage for detecting SNV when using WES compared to whole genome sequencing (e.g. 100x vs 15x).

One merely requires per position read depth of ~30 in order to achieve high sensitivity to the gold standard when calling SNVs / SNPs. For somatic variants, however, high read depth is more desirable due to the fact that, in a tumour bulk biopsy, we would expect many somatic variants to only be detected at low frequencies.

Kevin

ADD COMMENTlink modified 5 weeks ago • written 5 weeks ago by Kevin Blighe61k
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