Go Back   SEQanswers > Applications Forums > Genomic Resequencing

Similar Threads
Thread Thread Starter Forum Replies Last Post
Reliability of short read annotations vanillasky Bioinformatics 0 05-17-2015 12:21 PM
insert sizes for RNA-seq luc RNA Sequencing 6 08-13-2014 11:07 AM
What insert size for velveth (1.2.10) with 2 sets of reads with diff. insert sizes? Genomics101 Bioinformatics 4 02-07-2014 12:41 PM
insert sizes nozzer Bioinformatics 1 07-09-2010 06:49 AM

Thread Tools
Old 05-14-2019, 01:57 AM   #1
Junior Member
Location: Newmarket, UK

Join Date: Apr 2017
Posts: 1
Default Reliability of short insert sizes

I have recently sequenced a number of PCR amplicons on an Illumina MiSeq, in a sample that must be a compund heterozygote for 2 SNPs (based on phenotype). The 2 SNPs are very close together, so in theory any read that covers both SNPs should have one of the variants, but never both. However, when I physically count them, only around 80% of reads follow this pattern, while the remaining 20% look like they have both variants, or neither.

The read length was set to 150bp, but quite a few of the reads (in the 20% group that don't appear as expected) are much shorter than that, as short as 40bp (when BAM files are viewed in IGV). Also, in many of the reads from the 20% group, one or both of the SNPs are within about 5 nucleotides of the end of the read.

I have since realised that the short reads are due to very short fragment/insert sizes to begin with (so the mate pairs are the same sequence/overlap 100%).

However, will this have an effect on the reliability of the variant calls?
If not, how do I explain that only 80% of the reads that span both SNPs (around 200 reads), have genotypes concordant with a compound heterozygote?
lburmz is offline   Reply With Quote

Thread Tools

Posting Rules
You may not post new threads
You may not post replies
You may not post attachments
You may not edit your posts

BB code is On
Smilies are On
[IMG] code is On
HTML code is Off

All times are GMT -8. The time now is 02:50 PM.

Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2019, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO