SEQanswers

Go Back   SEQanswers > Bioinformatics > Bioinformatics



Similar Threads
Thread Thread Starter Forum Replies Last Post
kmer counting software issue Rui Guo Bioinformatics 4 05-26-2016 01:26 AM
help with trinity and jellyfish bongbimit Bioinformatics 2 05-13-2015 02:01 AM
FastQC, Kmer count, Trimmomatic: no success in trimming, still fail Kmer skmotay RNA Sequencing 6 10-09-2014 06:24 AM
Jellyfish help bioman1 Bioinformatics 0 04-02-2014 12:52 AM
Trying to estimate genome size - no peak in jellyfish kmer abundance plot kmkocot Bioinformatics 5 03-27-2014 07:24 AM

Reply
 
Thread Tools
Old 02-11-2017, 01:20 AM   #1
Bourney
Junior Member
 
Location: South England

Join Date: Oct 2014
Posts: 5
Default Jellyfish kmer counting diploid

Hi there,

I've used jellyfish kmer count to estimate the genome size of a diploid organism. I get double peaks, which are expected, and the first one is twice the size, which is also expected when sequencing diploid. However, I've always been told the protocol is to disregard the first one, as it represents the heterozygous alleles, and use the peak of the second one for genome size estimation. But this advice has mainly been for haploid sequencing.
And this paper, https://academic.oup.com/dnaresearch...inbred-line-of, looking at a tetraploid, used the first peak in their estimation.

Would I be justified in using the first peak for estimation of a diploid genome? I can't think why I wouldn't, but don't want to have overlooked something critical.

For what it's worth, when using the first peak, I get an estimation of 170 Mbp, which is around the right size (same as sister-species), and when using SOAPdenovo and Abyss to assemble, I get an assembly twice that at 330-340 Mbp. I'm assuming that the assemblers aren't handling the diploidy overly well. So, as an aside, does anyone have any advice on making the assemblers incorporate the heterozygosity of a diploid genome?

Cheers
Attached Files
File Type: pdf fastqcounts.histo.pdf (12.8 KB, 30 views)
Bourney is offline   Reply With Quote
Old 02-11-2017, 09:33 AM   #2
Brian Bushnell
Super Moderator
 
Location: Walnut Creek, CA

Join Date: Jan 2014
Posts: 2,707
Default

The correct way to calculate the (haploid) genome size of a diploid is:

0.5*(# kmers in first [heterozygous] peak)+
1*(#kmers in second [homozygous] peak)+
2*(#kmers in third [2-copy] peak)+
...etc

Normally you can ignore the high peaks as they become increasingly hard to distinguish don't contribute a lot. But you absolutely can't disregard the first peak; in highly heterozygous organisms, that can constitute the bulk of the genome!

BBMap's KmerCountExact will generate a peaks file that does this automatically, when the peaks are clear (as in your example):

Quote:
kmercountexact.sh in=reads.fq khist=khist.txt peaks=peaks.txt ploidy=2
A header line in peaks.txt will indicate genome size (and calculated ploidy, if you don't specify it).
Brian Bushnell is offline   Reply With Quote
Reply

Tags
genome size estimation, jellyfish, kmer counting

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 11:12 AM.


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