SEQanswers

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



Similar Threads
Thread Thread Starter Forum Replies Last Post
SureSelect QXT: Transposons and 90-minute hyb ECO Sample Prep / Library Generation 1 08-02-2015 12:25 PM
PCR program for Sureselect libraries hawaii454-0 Illumina/Solexa 0 08-07-2012 07:51 AM
Evenness of coverage in RNA-seq library prep microphobe Sample Prep / Library Generation 0 04-04-2012 10:58 PM
Agilent SureSelect - coverage of high GC regions NGSfan Genomic Resequencing 28 11-07-2011 01:13 PM
SureSelect and Rapid MID Libraries DoubleA 454 Pyrosequencing 2 09-15-2010 02:05 PM

Reply
 
Thread Tools
Old 05-02-2017, 04:28 PM   #1
ngseq
Member
 
Location: Philadelphia, PA

Join Date: Mar 2010
Posts: 19
Unhappy Reduced coverage evenness of SureSelect QXT libraries with automation

Dear all,

We are trying to automate SureSelect QXT library prep with the Hamilton Star liquid handler, and encountered the issue of reduced coverage evenness. With automation, we had more coverage gaps even when the overall average coverage across the region of interest is slightly higher than the that of a manually prepped library. We are a clinical lab and coverage gaps have to be filled in by Sanger, which means that we would have to do more Sanger with automation of QXT library prep .

Does anyone have similar experience?

I suspect the following possible reasons:

1. Incomplete MyOne beads transfer and loss of captured DNA because of this. I heard that lower input DNA may actually lead to poor coverage evenness. So maybe MyOne beads loss has similar effect?

2. On-deck hybridization: evaporation and volume loss during hybridization set up. The liquid handler takes 3-5 minutes to set up the hybridization on the deck, so I am sure there was some volume loss due to evaporation.

Any thoughts or comments are much appreciated. thanks!
ngseq is offline   Reply With Quote
Reply

Tags
uneven qxt coverage

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 06:47 AM.


Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2020, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO