Go Back   SEQanswers > Bioinformatics > Bioinformatics

Similar Threads
Thread Thread Starter Forum Replies Last Post
Low frequency variant caller for any ploidy level me_myself_andI Bioinformatics 16 04-21-2014 08:39 AM
filter low complexity with GATK matan8 Bioinformatics 0 02-27-2012 03:00 AM
allele frequency threshold GATK m_elena_bioinfo Introductions 3 12-12-2011 12:29 PM
PubMed: Resequencing of 200 human exomes identifies an excess of low-frequency non-sy Newsbot! Literature Watch 1 12-02-2010 08:04 AM
samtools pileup for low frequency alleles? greigite Bioinformatics 0 03-02-2010 04:43 PM

Thread Tools
Old 05-17-2012, 02:26 PM   #1
Junior Member
Location: Boston

Join Date: Jan 2011
Posts: 9
Default Detecting low frequency mutations - GATK

I am analyzing a targeted enrichment of a Hapmap blend. I have 4 hapmap samples mixed in the ratio: 74:20:5:1
I am looking to detect variants at low frequencies (upto 0.5%).
I am using GATK for the SNP analysis - I carried out sample level local realignment around indels and base quality score recalibration.
With Unified Genotyper, I am getting a very good expected: actual ratio for SNPs above 10% expected frequency. However, I am not picking up any of the SNPs in the 0.5 to 10% range. I do see a lot of these SNPs (on CLC bio Genomics workbench) and pick them up the CLC bio SNP caller (along with 100's of false positives). I understand a lot of them may be poor quality or may have been rejected by GATK due to strand bias or other problems. But the fact that not one of the 20 expected calls were picked up makes me wonder if I am doing something wrong.
The coverage in these positions is very high (~2000). Sequencing was done on Illumina. I used the default quality score settings on GATK. Is there any parameter that I can work around to increase the sensitivity on this particular set and pick up these low frequency SNPs ?
Any help would be appreciated.
spreeth84 is offline   Reply With Quote
Old 02-11-2015, 06:38 AM   #2
Junior Member
Location: Sweden

Join Date: Sep 2010
Posts: 2
Default Low frequency SNPs with GATK?

I found this post and have about the same question as spreeth84 so I will send this and see if this thread can come to life.

I have 30 small haploid bacterial genomes where I would like to find SNPs at low frequency. Ive used the HaploType-caller

java -jar GenomeAnalysisTK.jar -T HaplotypeCaller -R reference.fna -I S1.bam -I S2.bam -I S3.bam -I S4.bam .... -I S30.bam -ploidy 1 -stand_call_conf 30 -stand_emit_conf 10 -o variants.vcf -dcov 1000000000
Im using the GenomeAnalysisTK-3.3-0-g37228af version of GATK.

Im doing multiple sample calling and I see SNPs with lower frequencies (say 20%) but only when some other sample have a higher frequency at the same position.

Can I get all SNPs, not only high frequency ones with GATK, or should I use some other software?
swe is offline   Reply With Quote
Old 02-11-2015, 06:59 AM   #3
Junior Member
Location: Boston

Join Date: Jan 2011
Posts: 9

Hi swe, I have had better luck with Mutect than Haplotype caller or Unified Genotyper for calling low frequency alterations. There are parameters that can be further tweaked in Mutect to achieve higher sensitivity at low minor allele frequencies. I have never worked with Haploid bacterial genomes, so I don't know if there is any other reason this wouldn't work for you - but may be worth a shot.
spreeth84 is offline   Reply With Quote
Old 02-13-2015, 04:26 AM   #4
Senior Member
Location: Amsterdam

Join Date: Nov 2009
Posts: 112

We also tried Mutect and it indeed calls more true low frequency mutations.

@spreeth84: What parameters do you tweak? Do you know if there is a proper manual?
Zaag is offline   Reply With Quote

gatk, low frequency snp, sensitivity

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 08:18 PM.

Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2021, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO