VarScan.v2.4.1.description.txt

RELEASE NOTES FOR VARSCAN V2.4.1

23-Oct-2015

VarScan v2.4.1 is the current release
VarScan v2.4.0 was the first release to VarScan's new home at GitHub, http://dkoboldt.github.io/varscan/
VarScan v2.3.9 and prior releases will persist on SourceForge, as will the support forums and other resources. 

LICENSE
VarScan 2 is free for non-commercial use by academic, government, and non-profit/not-for-profit institutions. 
A commercial version of the software is available, and licensed through the Office of Technology Management at 
Washington University School of Medicine. For more information, please contact:

Paul Carter, Business Development Director
paulcarter@wustl.edu
+1 314-362-5426

VERSION 2.4.1 CHANGES
Major changes include:

1.) Correction of a bug in v2.4.0 that caused some somatic mutation calling oddities, especially for indels.
This issue was introduced in v2.4.0 (minor fix #1) in an attempt to more accurately count variant-supporting 
reads in the normal sample during somatic mutation calling. That fix is now rolled back. 

2.) Expansion and improvement of the fpfilter command. It now includes many more parameters for fine-tuning
the filter, and a master option (--dream3-settings 1) that will optimize parameters based on the DREAM-3 
mutation calling challenge (see below).

3.) The fpfilter command now outputs the average trimmed read length of reference- and variant-supporting 
reads, so there will be two new columns appended to the tab-delimited output file.


REMINDER: PLEASE USE THE FALSE POSITIVE FILTER
The scientific basis of this filter is described in the VarScan 2 publication. It will improve
the precision of variant and mutation calling by removing artifacts associated with short-read alignment.
-For somatic mutations, generate bam-readcounts with the Tumor BAM. For LOH and Germline, generate readcounts with the Normal BAM
-For de novo mutations (trio calling), generate readcounts with the child BAM.
The filter requires the bam-readcount utility: https://github.com/genome/bam-readcount

USAGE: java -jar VarScan.jar fpfilter [variant file] [readcount file] OPTIONS
	variant file - A file of SNPs or indels in VarScan-native or VCF format
	readcount file - The output file from bam-readcount for those positions
	***For detailed filtering instructions, please visit http://varscan.sourceforge.net***

	OPTIONS:
	--output-file		Optional output file for filter-pass variants
	--filtered-file		Optional output file for filter-fail variants
	--dream3-settings	If set to 1, optimizes filter parameters based on TCGA-ICGC DREAM-3 SNV Challenge results
	--keep-failures		If set to 1, includes failures in the output file

	FILTERING PARAMETERS:
	--min-var-count		Minimum number of variant-supporting reads [4]
	--min-var-count-lc	Minimum number of variant-supporting reads when depth below somaticPdepth [2]
	--min-var-freq		Minimum variant allele frequency [0.05]
	--max-somatic-p		Maximum somatic p-value [0.05]
	--max-somatic-p-depth	Depth required to test max somatic p-value [10]
	--min-ref-readpos	Minimum average read position of ref-supporting reads [0.1]
	--min-var-readpos	Minimum average read position of var-supporting reads [0.1]
	--min-ref-dist3		Minimum average distance to effective 3' end (ref) [0.1]
	--min-var-dist3		Minimum average distance to effective 3' end (var) [0.1]
	--min-strandedness	Minimum fraction of variant reads from each strand [0.01]
	--min-strand-reads	Minimum allele depth required to perform the strand tests [5]
	--min-ref-basequal	Minimum average base quality for ref allele [15]
	--min-var-basequal	Minimum average base quality for var allele [15]
	--min-ref-avgrl		Minimum average trimmed read length for ref allele [90]
	--min-var-avgrl		Minimum average trimmed read length for var allele [90]
	--max-rl-diff		Maximum average relative read length difference (ref - var) [0.25]
	--max-ref-mmqs		Maximum mismatch quality sum of reference-supporting reads [100]
	--max-var-mmqs		Maximum mismatch quality sum of variant-supporting reads [100]
	--max-mmqs-diff		Maximum average mismatch quality sum (var - ref) [50]
	--min-ref-mapqual	Minimum average mapping quality for ref allele [15]
	--min-var-mapqual	Minimum average mapping quality for var allele [15]
	--max-mapqual-diff	Maximum average mapping quality (ref - var) [50]


DREAM-3 SETTINGS FOR FPFILTER
Please note the --dream3-settings parameter for fpfilter, which (if set to 1) will optimize the 
false positive filter settings based on the fine-tuning we did for the TCGA-ICGC DREAM-3 
SNV Challenge. See the "in silico 3" dataset described here:
https://www.synapse.org/#!Synapse:syn312572/wiki/62018
This dataset modeled 100% tumor purity, but three subclones at 50%, 33%, and 20% variant allele frequency.
Optimal VarScan settings were established as follows:
	For SAMtools:
		mpileup -B (disables BAQ)

	For VarScan somatic:
		--min-coverage 3 --min-var-freq 0.08 --p-value 0.10 --somatic-p-value 0.05 --strand-filter 0

	For VarScan fpfilter:
		--min-var-count = 3
		--min-var-count-lc = 1
		--min-strandedness = 0
		--min-var-basequal = 30
		--min-ref-readpos = 0.20
		--min-ref-dist3 = 0.20
		--min-var-readpos = 0.15
		--min-var-dist3 = 0.15
		--max-rl-diff = 0.05
		--max-mapqual-diff = 10
		--min-ref-mapqual = 20
		--min-var-mapqual = 30
		--max-var-mmqs = 100
		--max-ref-mmqs = 50

CITING VARSCAN
If you use VarScan, please note the version number and cite this publication along with the 
version-appropriate URL:

Koboldt DC, Zhang Q, Larson DE, Shen D, McLellan MD, Lin L, Miller CA, Mardis ER, Ding L, Wilson RK. 
VarScan 2: somatic mutation and copy number alteration discovery in cancer by exome sequencing. 
Genome Res. 2012 Mar;22(3):568-76. doi: 10.1101/gr.129684.111. 

https://github.com/dkoboldt/varscan (v2.4.0 and beyond)
or
http://varscan.sourceforge.net (v2.3.9 and before)