Question: exome sequencing, target capture
gravatar for tonja.r
4.3 years ago by
tonja.r450 wrote:

I have some problems with the understanding of the part of exome sequencing where the targets (exons) are captured.  And how the SNPs (or SNVs) can be detected.

Firstly, the genomic DNA is shared, so we have exomes, introns there and intergeneric regions there. The primers are attached and then the hybridization takes place which helps to get rid of non-exome sequences. It means that we need to know the sequences of exons in forehand. So, I need to design exon sequences that already include all possible SNPs and due to those sequences I can capture my patient's exons with SNPs, that I know in forehand. If some sequence do not hybridize, it means there is no SNP which is associated with this sequence in my patient. How can I detect then the novel SNPs or SNVs in exome sequencing if for capturing the targets I had to know the corresponding SNPs and SNVs?

Thank you in advance.

sequencing snp exome • 1.6k views
ADD COMMENTlink modified 4.3 years ago by Brian Bushnell16k • written 4.3 years ago by tonja.r450
gravatar for Brian Bushnell
4.3 years ago by
Walnut Creek, USA
Brian Bushnell16k wrote:

Commercial exon capture kits for NGS do not require exact matches; they hybridize with sequences that have SNPs, indels, etc.  I don't know exactly how low an identity will still be captured, but a few SNPs, or even long indels, don't seem to cause any problems.  You then determine which variations are present by mapping the reads to a reference.

If you are designing your own primers to specifically amplify a certain region, then indeed you might run into problems if there are mutations in the primer regions.

ADD COMMENTlink written 4.3 years ago by Brian Bushnell16k
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