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Forum: Bioinformatics 09-03-2013, 09:18 AM
Replies: 3
Views: 2,654
Posted By dkoboldt
Thank you for posting. From the VarScan VCF entry...

Thank you for posting. From the VarScan VCF entry for sample 2:
0/0:32:55:37:32:5:13.51%:2.706E-2:38:15:15:17:3:2

You can see that the VAF as computed by VarScan is 13.51%, which is below your...
Forum: Bioinformatics 09-03-2013, 09:11 AM
Replies: 11
Views: 3,828
Posted By dkoboldt
Hello, and thanks for posting. Do you mean that...

Hello, and thanks for posting. Do you mean that the VCF header lines are missing? That's strange behavior and I'm happy to help you investigate.

Would you mind letting me know what your output...
Forum: Bioinformatics 09-03-2013, 08:33 AM
Replies: 1
Views: 3,177
Posted By dkoboldt
Are there situations where true, validated...

Are there situations where true, validated somatic mutations arise at the position of heterozygous SNPs? My expectation is that most such situations would represent artifactual calls, not real...
Forum: Bioinformatics 09-03-2013, 08:25 AM
Replies: 1
Views: 2,409
Posted By dkoboldt
This issue is caused by the -u parameter given to...

This issue is caused by the -u parameter given to SAMtools. Remove -u and it should work fine.
Forum: Bioinformatics 09-03-2013, 08:20 AM
Replies: 11
Views: 6,213
Posted By dkoboldt
Thanks for letting me know!

Thanks for letting me know!
Forum: Bioinformatics 08-06-2013, 12:53 PM
Replies: 10
Views: 4,595
Posted By dkoboldt
Hello, and thanks for posting this issue. Yes,...

Hello, and thanks for posting this issue. Yes, VarScan does not expect to see a line with coverage=0 in a single-sample pileup file. VarScan v2.3.6 addresses this issue and should not crash. In...
Forum: Bioinformatics 06-28-2013, 01:56 PM
Replies: 4
Views: 5,422
Posted By dkoboldt
Elena, You could probably just perform steps...

Elena,

You could probably just perform steps 1 and 3 and get a good high-confidence set of somatic mutation calls.... the somaticFilter command is more simplistic than the filter false positives. ...
Forum: Bioinformatics 03-20-2013, 03:47 PM
Replies: 11
Views: 6,213
Posted By dkoboldt
Hello all, We have just released VarScan...

Hello all,

We have just released VarScan v2.3.5 which should correct the invalid mpileup warning:

https://sourceforge.net/projects/varscan/files/
Forum: Bioinformatics 03-19-2013, 11:21 AM
Replies: 1
Views: 2,342
Posted By dkoboldt
Hello, and thank you for your question. When you...

Hello, and thank you for your question. When you specify VCF output format, VarScan reports sites that failed its internal filters, such as the strand-filter, and this is indicated in the FILTER...
Forum: Bioinformatics 03-19-2013, 09:19 AM
Replies: 11
Views: 6,213
Posted By dkoboldt
Vyellapa, can you send me the first 75 lines of...

Vyellapa, can you send me the first 75 lines of your pileup file? Send it to dkoboldt (at) genome [dot] wustl [dot] edu
Forum: Bioinformatics 01-30-2013, 10:30 AM
Replies: 22
Views: 12,979
Posted By dkoboldt
Hello all, Thank you for following up on...

Hello all,

Thank you for following up on this issue -- we certainly do want VarScan's VCF output to meet the specification. I believe that I've already addressed the missing QUAL field, and the...
Forum: Bioinformatics 01-30-2013, 10:03 AM
Replies: 11
Views: 6,213
Posted By dkoboldt
It's strange to suddenly encounter an...

It's strange to suddenly encounter an invalid-format line in SAMtools mpileup output. If I take it *exactly* as you pasted it, there's no delimiter (tab) between the last base call in column 5 and...
Forum: Bioinformatics 01-30-2013, 09:52 AM
Replies: 3
Views: 1,096
Posted By dkoboldt
Thanks for the question, rdeboja, and to adrianl...

Thanks for the question, rdeboja, and to adrianl for the answer. I will make sure the output is properly headered in the next release of VarScan!
Forum: Bioinformatics 01-30-2013, 09:50 AM
Replies: 2
Views: 1,092
Posted By dkoboldt
Jackken, That's a bit puzzling... it looks...

Jackken,

That's a bit puzzling... it looks like you didn't provide a reference to SAMtools mpileup (-f) when generating the pileup file, which is why the reference base is always "N". This can...
Forum: Bioinformatics 01-30-2013, 09:37 AM
Replies: 2
Views: 2,234
Posted By dkoboldt
John, I would restructure your command...

John,

I would restructure your command slightly so that redirecting the output is the last thing you do... VarScan might not be interpreting your "--mpileup 1" :
samtools mpileup -q 1 -f...
Forum: Bioinformatics 01-30-2013, 09:27 AM
Replies: 4
Views: 3,895
Posted By dkoboldt
Chris, I'm glad you came across the solution...

Chris,

I'm glad you came across the solution - it looks like there's a universal issue mapping reads in "proper" pairs at that location. Thanks for using VarScan!
Forum: Bioinformatics 01-30-2013, 08:59 AM
Replies: 1
Views: 1,284
Posted By dkoboldt
Ng, I think that happened because you used the...

Ng, I think that happened because you used the "--variants" flag, which overrides your command (mpileup2snp) and outputs both SNPs and indels. Remove that flag and you should be good to go.
Forum: Bioinformatics 12-27-2012, 08:49 AM
Replies: 1
Views: 1,888
Posted By dkoboldt
Hello, Those are heterozygous genotypes...

Hello,

Those are heterozygous genotypes represented as IUPAC ambiguity codes (http://droog.gs.washington.edu/parc/images/iupac.html).
Forum: Bioinformatics 12-07-2012, 12:37 PM
Replies: 3
Views: 1,949
Posted By dkoboldt
I strongly recommend updating to the latest...

I strongly recommend updating to the latest release of VarScan (http://sourceforge.net/projects/varscan/files/?source=navbar) (v2.3.3), which outputs more extensive information into VCF format...
Forum: Bioinformatics 12-07-2012, 12:35 PM
Replies: 2
Views: 3,168
Posted By dkoboldt
A couple of cautions when doing variant calling...

A couple of cautions when doing variant calling using RNA-seq data:
1.) False positive rates are higher, both due to RT errors and the extra alignment difficulty of mapping mature mRNA transcripts...
Forum: Bioinformatics 11-08-2012, 10:45 AM
Replies: 4
Views: 2,464
Posted By dkoboldt
Thank you again for following up. I'd thought...

Thank you again for following up. I'd thought that the locale-parsing issue was fixed, but using your file was able to duplicate the problem and fix it. Now VarScan copyCaller should work for you...
Forum: Bioinformatics 11-08-2012, 09:13 AM
Replies: 5
Views: 1,670
Posted By dkoboldt
Agreed, make the reference chromosome names as...

Agreed, make the reference chromosome names as simple as possible. Make sure the reference that you aligned to is the same you provide for mpileup. If you still encounter the floating point...
Forum: Bioinformatics 11-08-2012, 08:58 AM
Replies: 7
Views: 2,829
Posted By dkoboldt
I'm hereby promoting Jane M to "VarScan veteran"....

I'm hereby promoting Jane M to "VarScan veteran". Thanks for the answers!
Forum: Bioinformatics 11-08-2012, 08:56 AM
Replies: 3
Views: 2,036
Posted By dkoboldt
As the others said, use mpileup but be aware that...

As the others said, use mpileup but be aware that there are differences between the two commands, most notably that BAQ adjustment is turned on by default in mpileup, which affects base qualities....
Forum: Bioinformatics 11-08-2012, 08:42 AM
Replies: 2
Views: 1,965
Posted By dkoboldt
This is a parsing error, usually encountered when...

This is a parsing error, usually encountered when VarScan is given an input file that's not the expected format. Can you provide your full command, as well as an excerpt from each of your input...
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