SEQanswers

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



Similar Threads
Thread Thread Starter Forum Replies Last Post
ATAC-seq primers Fasteno Sample Prep / Library Generation 13 03-12-2019 10:12 PM
ATAC-seq Laiumiunix Sample Prep / Library Generation 0 04-03-2018 03:35 PM
On-Plate ATAC-seq krutipedali Sample Prep / Library Generation 2 02-28-2017 04:16 AM
ATAC-seq QC using gel Fasteno Sample Prep / Library Generation 0 03-12-2015 04:13 AM

Reply
 
Thread Tools
Old 03-13-2019, 01:06 AM   #1
pritb
Junior Member
 
Location: Singapore

Join Date: Mar 2019
Posts: 2
Unhappy Need help in ATAC-seq

Hello All

I am new to ATAC seq experiments. I am working on human fibroblast with 50k cells (counted through countess). Initially, I started following J Buenrostro (2015) protocol but seems not working. Now I shifted myself to Omni-ATAC protocol.

My Initial OD (after Tn5 treatment) using Nanodrop gave 46 ng/ul. I proceeded with Library prep by giving 5 cycles. Realtime PCR for library quantification gave me 20 additional cycles for my Libraries. After PCR cleanup, I got 96 ng/ul as library concentration. On gel check, I cant seen anything, except for few bands that is less than 100 bps. What could be reason?

I have attached my gel picture here.
Lane-1: 2 ul of sample after Tn5 reaction post cleanup
Lane-2: 15 ul of sample after 35 cycles (Although I selected 20 cycles based on 1/4 fluorescence intensity)
Lane-3: 10 ul of sample of 20 cycles (additional PCR cycles). This is purified. You can seen 2 bands less than 100 bps.

Can anyone help me out?
Attached Images
File Type: jpg Presentation1.jpg (37.5 KB, 8 views)
pritb 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 01:45 PM.


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