Go Back   SEQanswers > Sequencing Technologies/Companies > Illumina/Solexa

Similar Threads
Thread Thread Starter Forum Replies Last Post
Insert size important? 454andSolid De novo discovery 3 12-27-2017 01:33 AM
How to get library size sajoshi Sample Prep / Library Generation 2 05-12-2016 03:13 PM
HTSeq version in-consistency and the STRANDED option LeonDK Bioinformatics 4 09-18-2014 08:57 PM
qPCR consistency hawaii454-0 Sample Prep / Library Generation 1 07-20-2012 10:16 AM

Thread Tools
Old 08-07-2017, 02:55 AM   #1
Junior Member
Location: UK

Join Date: Dec 2015
Posts: 2
Default Is the consistency of the NGS library size important?

Hello everyone! I have a general question about the importance of keeping the library fragment size consistent between samples. All my libraries have fragments ranging from 200 to 1000 bp according to the Bioanalyzer, which is fine for the Illumina paired-end sequencing. However, some of the libraries have average size of let's say 300 bp, others - 350-400 bp. I was wondering if this could be a problem by introducing some kind of bias? As far as I can see as long as the alignment rate is the same between the different samples, there shouldn't be any other hidden source of bias. In my particular example, I am doing spiked-in RNA-seq and ChIP-seq experiments and I need to prepare libraries from sonicated genomic DNA to find out the ratio of drosophila to mouse reads in the sample to determine the consistency of the spike-in. For some reason, the sonication efficiency varies slightly from sample to sample, which results in the libraries that have slightly different fragment size distribution. I would really appreciate if you could share with me your opinion/experience/piece of advance on this matter!
mrshope is offline   Reply With Quote
Old 08-07-2017, 03:28 AM   #2
Senior Member
Location: East Coast USA

Join Date: Feb 2008
Posts: 6,787

In practice smaller fragments always cluster more efficiently. So you should keep that in mind. As long as your library has fragments predominantly in the 300-400 bp range this should not be a big issue but if most fragments are ~1kb then you may be biasing away from those.
GenoMax 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 06:48 PM.

Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2018, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO