This is a test version of Biostars. For the public version, visit https://www.biostars.org.
Input Error Using Varscan2.3

Hi,

I'm using Varscan2.3 to generate vcf files. I understand that we need to convert the BAM files to mpileup format first and then give it as an input. But I keep getting an error.

The command I used:

samtools mpileup -f ~/refs/human_g1k_v37.fasta file.bam | java -jar /varscan/VarScan.v2.3 mpileup2snp --output-vcf > raw.vcf

I get the following error:

Error: Invalid format for pileup at line 12
1       11829   A       0

There was a thread already (Varscan Error On Mpileup Input), but that didn't help me. Does anyone know how to resolve this?

Thanks!

error mpileup
• 5,068 views • link
updated by dankoboldt • dankoboldt • written by Jordan Jordan

Hi, I wonder if you have solved the problem, if so, would you mind giving me some suggestions on how to solve this?

• 0 views • link
written by Liye Zhang Liye Zhang

I did actually. I don't remember the exact command but, what I did was instead of piping it, I created mpileup's for both tumor and normal first. And then I used this command on the mpileup's

java -jar VarScan.jar somatic normal.pileup tumor.pileup --mpileup 1

This gives you the somatic variants directly.

• 0 views • link
updated by Ram Ram • written by Jordan Jordan

2 answers

This is an issue due to the zero-depth lines reported by SAMtools mpileup, which VarScan does not expect, and should be fixed in v2.3.6. Please let me know if it's still an issue!

• 526 views • link
updated by Ram Ram • written by dankoboldt • dankoboldt

I resolved the problem, but this is something you might want to know.

I tried the samtools mpileup and pipe to Varscan.

It did not work when I used samtools 0.1.19 for Varscan2.2.11 ,Varscan2.3.5 and Varscan2.3.6.

But if I use the samtools 0.1.18, all versions worked.

Conclusion: the newer version of samtools may cause the problem.

• 0 views • link
updated by Ram Ram • written by Liye Zhang Liye Zhang

Try using pileup2snp instead of mpileup2snp (even if you use mpileup to build the pileup). This usually works for me.

• 1 views • link
updated by Ram Ram • written by Fabio Marroni Fabio Marroni

Log in to answer this question.

More posts like this
  • Samtools Mpileup Does Not Contain Sample Name
    written by komal.rathi komal.rathi
    <p>Hi everyone,</p> <p>I am using <a href='http://samtools.sourceforge.net/'>samtools</a> mpileup to generate a mpileup file from 64 <a href='http://samtools.sourceforge.net/SAM1.pdf'>bam</a> files using a set of chromosomal positions in …
  • Error While Running Cuffdiff On Multiple Bam Files Of A Group
    written by Jordan Jordan
    <p>Hi,</p> <p>I'm trying to run cuffdiff on 6 samples, divided into two groups of 3 samples each - G1(Female), G2(Male). But, it does not run. …
  • Samtools Mpileup Defined Regions
    written by Vova Naumov Vova Naumov
    <p>Hi! I need to get genotypes in a list of positions (for example BED file) having <a href='http://samtools.sourceforge.net/SAM1.pdf'>BAM</a> file. Trying to use tool <a href='http://samtools.sourceforge.net/'>samtools</a> …
  • Mpileup Problem
    written by stolarek.ir stolarek.ir
    <p>Hi.</p> <p>I have a problem with my variant calling using <a href='http://samtools.sourceforge.net/'>samtools</a>. I'm interested in vcf file with indels in my <a href='http://samtools.sourceforge.net/SAM1.pdf'>bam</a> file. </p> …
  • Why Is The Variant File Produced By Samtools So Large? It Is About ~34G.
    written by Jordan Jordan
    <p>Hi,</p> <p>I have a <a href='http://samtools.sourceforge.net/SAM1.pdf'>bam</a> file of size 62G. When I do the variant analysis by <a href='http://samtools.sourceforge.net/'>samtools</a>, it gives a huge bcf file …
  • Snp Calling With Bcftools Using Pileup File Format Instead Of Bcf
    written by Justin Justin
    <p>Usually when I do SNP calling, I do this:</p> <pre><code>samtools mpileup -g -f Ref.fa File.bam | bcftools view -cv - </code></pre> <p>where <a href='http://samtools.sourceforge.net/'>samtools</a> outputs …
  • Samtools Depth Stdin ?
    written by Mike Axtell Mike Axtell
    <p>Hi all,</p> <p>I would like to use <a href='http://samtools.sourceforge.net/'>samtools</a> depth in a pipeline to grab strand-specific coverage from a given interval, where the <a href='http://samtools.sourceforge.net/SAM1.pdf'>bam</a> …
  • Trouble Sorting A Bam File
    written by Jordan Jordan
    <p>Hi,</p> <p>I'm having trouble sorting a <a href='http://samtools.sourceforge.net/SAM1.pdf'>bam</a> file. I give the following command.</p> <pre><code>samtools sort -m 40G -o file.bam file.sorted </code></pre> <p>But I get …
  • Varscan Error On Mpileup Input
    written by GPR GPR
    <p>Hello, I am trying to filter low quality data off my mpileup files using Varscan but keep getting stuck with the error message: Invalid format …
  • Sam To Bam Conversion Reference Fasta File
    written by Atom Smasher • Atom Smasher
    <p>Hello,</p> <p>I had some data in "eland_export.txt" format and I want to convert it to the <a href='http://samtools.sourceforge.net/SAM1.pdf'>SAM</a> format first and then later to the …