SEQanswers

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



Similar Threads
Thread Thread Starter Forum Replies Last Post
ChIP-seq library prep Illumina Kapa ETHANol Sample Prep / Library Generation 9 01-29-2015 03:09 PM
increased bias in single-buffer library preps (NEB, KAPA,...) Reason for concern? luc Sample Prep / Library Generation 0 06-10-2014 09:49 AM
Has anyone tried comparing accuracy using different library preps? oiiio Bioinformatics 0 12-06-2013 08:04 AM
SPRI-TE Illumina library preps GW_OK Sample Prep / Library Generation 5 08-23-2010 09:13 AM
Ligase for DIY GAII library preps nickb Sample Prep / Library Generation 4 03-05-2010 10:36 AM

Reply
 
Thread Tools
Old 10-14-2014, 03:24 PM   #1
atcghelix
Member
 
Location: CA

Join Date: Jul 2013
Posts: 74
Default Stretching Kapa library preps

Hi Everyone,

Has anyone tried doing half reactions with the Kapa library prep kits using 1000ng of input DNA? I'm curious if the amount of reagents in a half-reaction is enough to ligate adapters to 1000ng of input DNA.

Thanks very much!
atcghelix is offline   Reply With Quote
Old 10-14-2014, 10:54 PM   #2
nucacidhunter
Jafar Jabbari
 
Location: Melbourne

Join Date: Jan 2013
Posts: 1,238
Default

If you are going to do lots of library preps, it might be worth to do a comparison yourself. Prepping two libraries from the same input DNA and using 1/2 reactions with one and full reaction with other and comparison of PCR yield without sequencing would give some indications on performance of 1/2 reaction. With TruSeq kit, 1/2 and 1/3 reactions works as good as full reactions. Generally most kits use excessive amount of reagents to ensure good performance even with poor laboratory practices and low quality input.
nucacidhunter is offline   Reply With Quote
Old 10-19-2014, 12:10 AM   #3
bilyl
Member
 
Location: USA

Join Date: Aug 2013
Posts: 52
Default

Quote:
Originally Posted by atcghelix View Post
Hi Everyone,

Has anyone tried doing half reactions with the Kapa library prep kits using 1000ng of input DNA? I'm curious if the amount of reagents in a half-reaction is enough to ligate adapters to 1000ng of input DNA.

Thanks very much!
I routinely shear a couple hundred ng of DNA with a Covaris then concentrate it down to 20ul with an Amicon Ultra-0.5 10k. From there I can scale down my Kapa Hyper Prep reagents to 0.4x of what they should be. Haven't seen any problems yet!
bilyl 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 01:08 AM.


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