Go Back   SEQanswers > Bioinformatics > Bioinformatics

Similar Threads
Thread Thread Starter Forum Replies Last Post
RPKMs for paired-end ATAC-seq data LacquerHead Bioinformatics 0 11-01-2016 07:22 AM
Haplotype phasing from Miseq 150bp paired end reads? michael-steffen Bioinformatics 3 01-08-2015 11:41 PM
RNA-Seq: Single Read and Paired End mRNA-Seq Illumina Libraries from 10 Nanograms Tot Newsbot! Literature Watch 0 11-09-2011 02:10 AM
Assembly of 150bp paired-end reads anli Illumina/Solexa 1 05-18-2011 03:01 AM

Thread Tools
Old 09-01-2017, 04:49 AM   #1
Location: canada

Join Date: Nov 2010
Posts: 20
Default Is it OK to use paired-end 150bp to sequence ATAC-seq libraries?


I am planning to do ATAC-seq assays and I am wondering if it is fine to use paired-end 150bp to sequence ATAC-seq libraries?
Just because the sequencing platform I am working with is only doing 150bp run with HiSeq3000...
I have the impression that people are usually using PE sequencing with 50 to 75bp reads. Would the only problem be that I would sequence a lot of adapters (exept money lost, adapter trimming should be enough to solve this)? Or would I lost many "small" fragments?

Thank you for your help!
vadoue is offline   Reply With Quote
Old 12-17-2017, 07:01 PM   #2
Junior Member
Location: Hong Kong

Join Date: Aug 2016
Posts: 2
Default Don't use large insert libraries for ATAC-seq


This is probably too late to be of any help to you, but for anyone else doing ATAC-seq, I highly recommend doing paired-end sequencing with 50x50bp or lower.

Even with adapter trimming, I have had a very difficult time mapping a 76x76bp PE Illumina NextSeq ATAC-seq library using bowtie2. There is always a dip in mapped reads around the 76bp insert size (see attached example plot), and this has subsequently caused NucleoATAC to have trouble generating its models for nucleosome positioning analysis.

And by have trouble, I mean the program crashes when running "nucleoatac run ..." making that analysis method unusable. There was an issue created in the NucleoATAC GitHub repo that discusses this problem a little bit (

Attached Images
File Type: jpeg example.jpeg (15.3 KB, 37 views)
cbp44 is offline   Reply With Quote
Old 12-21-2017, 06:30 AM   #3
Senior Member
Location: Purdue University, West Lafayette, Indiana

Join Date: Aug 2008
Posts: 2,317

You could always hard-clip your reads to 50 bases.

pmiguel is offline   Reply With Quote
Old 12-21-2017, 06:28 PM   #4
Junior Member
Location: Hong Kong

Join Date: Aug 2016
Posts: 2

Originally Posted by pmiguel View Post
You could always hard-clip your reads to 50 bases.
Hmm... that is something I hadn't thought of actually. I am going to give that a shot to see how the insert size distribution looks after that. I'll hard clip to 50 bases, then use trim_adapters ( to trim, but throw out reads smaller than 30bp after trimming.

I'd be interested if any other people had similar issues with an ATAC-seq library like this, and what solutions they would recommend.

I did try various adapter trimmers on the 76x76bp data and compared the mapping results. For example, I tried trim_adapters, trimmomatic and cutadapt both with and without the trimming done by bcl2fastq (so 6 different possibilities total). Using bcl2fastq trimming, and then trim_adapters seemed to work the best for me, but there wasn't much of a difference.

Most recently I tried remapping with very slow bowtie2 settings to see if that would help. For a Mus musculus sample with about 60 million paired-end reads, it took about 5 days to finish mapping (12 threads, ~2.6GHz per), but it did actually help a bit by getting me 8-10% more uniquely mapping concordant pairs, and more 70-80bp insert size reads to map.

For reference, these are the bowtie2 settings I used: "-k 4 --dovetail --no-mixed -D 20 -R 3 -N 1 -L 8 -i S,1,0.50". Mostly the same as "--very-sensitive-local", just lowering the seed size (-L), and allowing for a mis-match (-N 1).

--very-sensitive-local == -D 20 -R 3 -N 0 -L 20 -i S,1,0.50
cbp44 is offline   Reply With Quote

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 08:11 PM.

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