SEQanswers

Go Back   SEQanswers > Applications Forums > Sample Prep / Library Generation



Similar Threads
Thread Thread Starter Forum Replies Last Post
Library prep woes unusual bioanalyzer profile of Illumina RNA-seq library petey Sample Prep / Library Generation 1 11-19-2012 09:00 AM
Weird bioanalyser profile nisha barak Sample Prep / Library Generation 5 06-02-2012 08:41 AM
Weird Illumina library ashchin Illumina/Solexa 7 02-17-2012 11:34 AM
Weird library with ScriptSeq cascoamarillo Sample Prep / Library Generation 3 01-24-2012 06:58 AM
To Helicos or Not? Weird bioanalyzer profile: DNA or RNA? Dapip33 Sample Prep / Library Generation 6 02-04-2011 07:11 AM

Reply
 
Thread Tools
Old 03-11-2013, 02:24 AM   #1
JustinNelligan
Junior Member
 
Location: France

Join Date: Mar 2013
Posts: 4
Exclamation Weird library profile

Hello,

We are seeing a weird library profile - double peak + some high molecular weight smear after the range we cut. It's a chip-seq library amplified for 18 cycles - first time we see this specific profile, should we sequence it?

Thanks!

weird-library.jpg
JustinNelligan is offline   Reply With Quote
Old 03-25-2013, 09:21 AM   #2
kbusseq
Junior Member
 
Location: Tempe, AZ

Join Date: Mar 2013
Posts: 7
Default

It's possible that the double peak is caused by running too much sample for the chip (I'm guessing your image is from a Bioanalyzer chip or something similar). We've seen some seriously bizarre peaks on our Bioanalyzer (some double like yours and one that was actually a triple peak) that are remedied simply by re-running the sample at a 1:5 or 1:10 dilution.

Sometimes this eliminates all trace of a high MW smear as well, but typically I would guess that the presence of high MW fragments is due to over-amplifying the library. Do you usually amplify for 18 cycles? That is a little high in my experience (which, granted, I don't have tons of yet... ), but if it normally works for you than it might not be the issue.
kbusseq 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 10:49 AM.


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