SEQanswers

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



Similar Threads
Thread Thread Starter Forum Replies Last Post
TrueSeq PCR free vs Nextera Mitogenome seq frascom Sample Prep / Library Generation 4 05-23-2019 07:25 PM
NEB Fragmentase BTS Sample Prep / Library Generation 8 03-21-2014 10:43 AM
gDNA fragmentation NEBNext™ dsDNA Fragmentase™ jgibbons1 Illumina/Solexa 21 03-21-2014 08:01 AM
de novo assembling using NEBNext™ dsDNA Fragmentase™ hi-koike Illumina/Solexa 5 09-02-2013 01:42 PM

Reply
 
Thread Tools
Old 06-07-2019, 07:07 AM   #1
frascom
Junior Member
 
Location: Kansas

Join Date: May 2019
Posts: 4
Default NEB Fragmentase mitogenome advice

Hi, I am currently working with mitogenome sequencing of small mammals. I have amplified 5 PCR amplicons of the mitogenome around 4-5kb fragment lengths. My next step is to pool fragments and conduct shearing using NEB Fragmentase before library prep. Does anyone have any recommendations of the timing of incubation to achieve around 200bp fragments? My plan is to conduct a time series trial to find the optimum time, however, I am wary that low concentration of PCR Fragments that after shearing I may no actually be able to visualize on an agarose gel to confirm fragment shearing size. Conc of fragments ranges from 5ng/ul - 250 ng/ul.

Currently looking at end result using 300bp paired-end MiSeq kit for sequencing hence the 200bp fragment target.

But I am interested to know peoples experiences or recommendations to consider.

thanks for the help

Last edited by frascom; 06-07-2019 at 07:26 AM.
frascom is offline   Reply With Quote
Reply

Tags
fragment analysis, nebnext ultra, shearing

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 05:54 PM.


Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2019, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO