SEQanswers

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



Similar Threads
Thread Thread Starter Forum Replies Last Post
TruSeq PCR free DNA with 16 amplicons maddo1ck Illumina/Solexa 5 06-09-2016 02:07 AM
Doing PCR enrichment on PCR-free TruSeq lib? JBKri Sample Prep / Library Generation 2 10-08-2015 07:00 AM
TruSeq PCR-Free odd peaks aaronh Sample Prep / Library Generation 1 08-15-2014 08:30 PM
Nextera XT vs TruSeq PCR free vl80 Sample Prep / Library Generation 1 06-09-2014 01:13 AM
TruSeq PCR-free transition avo Sample Prep / Library Generation 2 03-05-2014 02:45 AM

Reply
 
Thread Tools
Old 01-11-2018, 05:08 AM   #1
vandykitty
Junior Member
 
Location: charlotte,nc

Join Date: Dec 2012
Posts: 7
Default TruSeq PCR Free question

I am preparing samples using the TruSeq DNA PCR Free library kit with UDI adaptors. The PI has requested that we perform Pippin Prep post adaptor ligation to remove adaptor contamination and target a narrow size distribution. Question is... because the protocol does not include denaturation prior to the final sequencing step, the non-homologous Y-shaped adaptors results in an atypical migration pattern on agarose gel and the Bioanalyzer results are substantially larger (750bp) than would be predicted (~350bp insert +adaptor range). Should I heat denature the samples prior to putting on Pippin Prep? Thoughts?

Last edited by vandykitty; 01-11-2018 at 07:40 AM.
vandykitty is offline   Reply With Quote
Old 02-19-2018, 12:36 PM   #2
aligenie
Member
 
Location: San Diego

Join Date: Feb 2011
Posts: 13
Default

Quote:
Originally Posted by vandykitty View Post
I am preparing samples using the TruSeq DNA PCR Free library kit with UDI adaptors. The PI has requested that we perform Pippin Prep post adaptor ligation to remove adaptor contamination and target a narrow size distribution. Question is... because the protocol does not include denaturation prior to the final sequencing step, the non-homologous Y-shaped adaptors results in an atypical migration pattern on agarose gel and the Bioanalyzer results are substantially larger (750bp) than would be predicted (~350bp insert +adaptor range). Should I heat denature the samples prior to putting on Pippin Prep? Thoughts?
I don't have any advice but want to know if you tried this and if it worked out.
aligenie is offline   Reply With Quote
Old 02-20-2018, 08:34 AM   #3
vandykitty
Junior Member
 
Location: charlotte,nc

Join Date: Dec 2012
Posts: 7
Default

The PI did not want to advert from the protocol without assurance from Illumina or Sage Science that this would work, so unfortunately, no we did not try this. Sending the samples out for sequencing today.
vandykitty 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 05:43 AM.


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