SEQanswers

Go Back   SEQanswers > Sequencing Technologies/Companies > Ion Torrent



Similar Threads
Thread Thread Starter Forum Replies Last Post
Coverage of reads not homogenous (sureselect target enrichement) Sam64 Genomic Resequencing 2 04-14-2017 09:37 AM
off-target reads cmccabe Bioinformatics 60 07-13-2013 06:39 AM
low sureselect on-target reads hawaii454-0 Sample Prep / Library Generation 3 04-10-2013 10:18 AM
Calculating number of reads per target base Lilach Bioinformatics 2 03-29-2012 09:37 AM
Where did my off target reads go? swNGS Bioinformatics 7 03-18-2012 12:26 PM

Reply
 
Thread Tools
Old 08-28-2013, 05:20 AM   #1
cmccabe
Senior Member
 
Location: chicago

Join Date: Jul 2012
Posts: 354
Default on-target reads

My on-target reads seem to be lower, about 80%, with this 86kb target panel. With our 35kb panel, the on-target reads were about 95%. We are using ampliseq and to make matters more complex when we barcode just 2 samples of the 86kb panel our on-target goes down to about 60%. Any sugesstions to improve the data? Thanks.
cmccabe is offline   Reply With Quote
Old 08-29-2013, 10:21 AM   #2
IonTorrent
Member
 
Location: Guilford, CT and S.F., CA

Join Date: Jan 2010
Posts: 64
Default

Quote:
Originally Posted by cmccabe View Post
My on-target reads seem to be lower, about 80%, with this 86kb target panel. With our 35kb panel, the on-target reads were about 95%. We are using ampliseq and to make matters more complex when we barcode just 2 samples of the 86kb panel our on-target goes down to about 60%. Any sugesstions to improve the data? Thanks.
Hi there,

In our experience, off-target reads are typically a combination of specific per panel and per sample effects. For example, the best HMW DNAs will tend to have very similar off-target rate, while FFPE DNA - particularly when very degraded, can yield higher off-target rates, and each panel will have an off-target contribution based on the specificity of the primers as well.

There is no reason why multiplexing two or more samples on one chip should be expected to increase off-target rate overall. If you are seeing an increase in off-target between samples with the same primer panel it could be sample specific. If you haven't done so already, I'd recommend a follow up with your Field Applications Scientist (FAS) to ensure there aren't other confounding factors that are at issue. They will be able to dig into the full set of data and offer guidance.
IonTorrent is offline   Reply With Quote
Old 08-29-2013, 10:27 AM   #3
cmccabe
Senior Member
 
Location: chicago

Join Date: Jul 2012
Posts: 354
Default on-target reads

I'm thinking maybe it is sample specific as for 6 runs the same pattern has been consistent. The NA12878 cell line has alsway given lower on-target reads (~50%), then the NS12911 (gDNA) sample which has an on-target ~86%. We are goig to try two gDNA samples to confirm. Thanks.
cmccabe is offline   Reply With Quote
Reply

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 01:10 AM.


Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2021, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO