-
Notifications
You must be signed in to change notification settings - Fork 4
/
Scons_variant_calling
439 lines (301 loc) · 26.1 KB
/
Scons_variant_calling
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
61
62
63
64
65
66
67
68
69
70
71
72
73
74
75
76
77
78
79
80
81
82
83
84
85
86
87
88
89
90
91
92
93
94
95
96
97
98
99
100
101
102
103
104
105
106
107
108
109
110
111
112
113
114
115
116
117
118
119
120
121
122
123
124
125
126
127
128
129
130
131
132
133
134
135
136
137
138
139
140
141
142
143
144
145
146
147
148
149
150
151
152
153
154
155
156
157
158
159
160
161
162
163
164
165
166
167
168
169
170
171
172
173
174
175
176
177
178
179
180
181
182
183
184
185
186
187
188
189
190
191
192
193
194
195
196
197
198
199
200
201
202
203
204
205
206
207
208
209
210
211
212
213
214
215
216
217
218
219
220
221
222
223
224
225
226
227
228
229
230
231
232
233
234
235
236
237
238
239
240
241
242
243
244
245
246
247
248
249
250
251
252
253
254
255
256
257
258
259
260
261
262
263
264
265
266
267
268
269
270
271
272
273
274
275
276
277
278
279
280
281
282
283
284
285
286
287
288
289
290
291
292
293
294
295
296
297
298
299
300
301
302
303
304
305
306
307
308
309
310
311
312
313
314
315
316
317
318
319
320
321
322
323
324
325
326
327
328
329
330
331
332
333
334
335
336
337
338
339
340
341
342
343
344
345
346
347
348
349
350
351
352
353
354
355
356
357
358
359
360
361
362
363
364
365
366
367
368
369
370
371
372
373
374
375
376
377
378
379
380
381
382
383
384
385
386
387
388
389
390
391
392
393
394
395
396
397
398
399
400
401
402
403
404
405
406
407
408
409
410
411
412
413
414
415
416
417
418
419
420
421
422
423
424
425
426
427
428
429
430
431
432
433
434
435
436
437
438
439
import os
#### Quick documentation ####
##Files needed by pipeline:
## - GRChxx.xx.fa
## - GRChxx.xx.fa.fai
## - GRChxx.xx.dict
## - dbSNP_vxxx_xxxx_noCHR.vcf - use remove_chr.py on vcf downloaded from dbSNP FTP site
## - Bed file containing exome captured regions - *.bed
## - config.py - python file containing variables to input in the pipeline. All variables must be declared before
#running the pipeline
##Program needed by pipeline:
## - picard.jar -
## - GenomeAnalysisTK.jar -
## - gatk - GATK4 for MuTect2
#########################
##### SCons Settings ####
#########################
vars = Variables("/working/configuration.py")
vars.Add('reference', 'The genome reference file',"reference.fa")
vars.Add('processors', 'Number of CPUs to be used', "2")
vars.Add('dbsnpVCF','The dbSNP VCF',"All_20180418.vcf.gz")
vars.Add('cosmic','The Cosmic VCF',"CosmicCodingMuts.vcf.gz")
vars.Add('exomeRegions',"The bed file containing exome regions","exome_regions.bed")
vars.Add('tumor',"Tumor sample","")
vars.Add('normal',"Normal sample","")
vars.Add('contamination','Computation of predicted contamination in tumor sample by CalculateContamination of GATK4 (y/n)',"n")
vars.Add('gatk4','The path to GATK4',"/tmp/gatk4/gatk")
vars.Add('javaOptions','Options to run java',"4")
vars.Add('maxPopulationAlleleFrequency',"Maximum population allele frequency of sites to consider for GetPileupSummaries in GATK4","0.2")
vars.Add('minPopulationAlleleFrequency',"Minimum population allele frequency of sites to consider for GetPileupSummaries in GATK4","0")
vars.Add('gnomadVCF',"The gnomad.exomes.rx.x.x.sites.pass.vcf","gnomad.exomes.r2.0.2.sites.vcf.gz")
vars.Add('variantCallers',"Variant callers to be used (mutect2,strelka2,varscan)","mutect2,strelka2,varscan")
vars.Add('gatk3','The path to gatk3',"/tmp/gatk3/GenomeAnalysisTK.jar")
# MuTect2 additional parameters
vars.Add('mutect2Parameters',"Parameters for MuTect2","")
# VarScan Parameters
vars.Add('varScanMinCoverage', 'VarScan minimum read depeh at a position to make a call [10]', "10")
vars.Add('varScanPvalue', "VarScan default p-value threshold for calling variants [1e-01]", "0.1")
vars.Add('varScanMinVarFreq', "Minimum variant allele frequency threshold [0.20]", "0.2")
vars.Add("varScanParameters", "Parameters to be added to varScan2", "")
# snpEff Parameters
vars.Add("snpeffDir", "SnpEFF directory", "/annotations")
vars.Add("snpeffGenomeVersion", "Genome version to be used by snpEff (like GRCh37.75)", "GRCh37.75")
# ClinVAr Parameter
vars.Add("clinvar", "Clinvar compressed VCF file", "clinvar_20190311.vcf.gz")
env = Environment(ENV = os.environ, SHELL = '/bin/bash', variables = vars)
env.AppendENVPath('PATH', os.getcwd())
Decider('timestamp-newer')
####################
##### Arguments ####
####################
reference = ARGUMENTS.get("reference", env["reference"])
reference = "/annotations/" + reference
processors = ARGUMENTS.get("processors", env["processors"])
dbsnpVCF = ARGUMENTS.get("dbsnpVCF",env["dbsnpVCF"])
dbsnpVCF = "/annotations/" + dbsnpVCF
cosmic = ARGUMENTS.get("cosmic",env["cosmic"])
cosmic = "/annotations/" + cosmic
exomeRegions = ARGUMENTS.get("exomeRegions",env["exomeRegions"])
exomeRegions = "/annotations/" + exomeRegions
clinvar = ARGUMENTS.get("clinvar",env["clinvar"])
clinvar = "/annotations/" + clinvar
tumor = ARGUMENTS.get("tumor",env["tumor"])
normal = ARGUMENTS.get("normal",env["normal"])
contamination = ARGUMENTS.get("contamination",env["contamination"])
gatk4 = ARGUMENTS.get("gatk4",env["gatk4"])
gatk3 = ARGUMENTS.get("gatk3", env["gatk3"])
javaOptions = ARGUMENTS.get("javaOptions",env["javaOptions"])
javaOptions = "-Xmx"+str(javaOptions)+"g"
maxPopulationAlleleFrequency = ARGUMENTS.get("maxPopulationAlleleFrequency",env["maxPopulationAlleleFrequency"])
minPopulationAlleleFrequency = ARGUMENTS.get("minPopulationAlleleFrequency",env["minPopulationAlleleFrequency"])
gnomadVCF = ARGUMENTS.get("gnomadVCF",env["gnomadVCF"])
gnomadVCF = "/annotations/" + gnomadVCF
variantCallers = ARGUMENTS.get("variantCallers",env["variantCallers"])
variantCallersList = variantCallers.split(",")
# MuTect2 additional parameters
mutect2Parameters = ARGUMENTS.get("mutect2Parameters",env["mutect2Parameters"])
# VarScan Parameters
varScanMinCoverage = ARGUMENTS.get("varScanMinCoverage", env["varScanMinCoverage"])
varScanPvalue = ARGUMENTS.get("varScanPvalue", env["varScanPvalue"])
varScanMinVarFreq = ARGUMENTS.get("varScanMinVarFreq", env["varScanMinVarFreq"])
varScanParameters = ARGUMENTS.get("varScanParameters", env["varScanParameters"])
# snpEff parameters
snpeffGenomeVersion = ARGUMENTS.get("snpeffGenomeVersion", env["snpeffGenomeVersion"])
#snpeffDir = "/annotations/" + snpeffGenomeVersion
snpeffDir = "/annotations/"
#snpeffDir = ARGUMENTS.get("sneffDir", env["snpeffDir"])
############################################
#### Calculate contamination with GATK4 ####
############################################
if contamination == "y" or contamination == "yes":
getPileupSummariesCMD = "${SOURCES[0]} GetPileupSummaries -I ${SOURCES[1]}/07_recalibrated.bam -L ${SOURCES[2]} -V ${SOURCES[3]} -O Variants/Mutect2/${SOURCES[1]}_${SOURCES[4]}/00_pileups.table" + " -max-af {} -min-af {}".format(maxPopulationAlleleFrequency,minPopulationAlleleFrequency)
getPileupSummaries = env.Command(["Variants/Mutect2/{}_{}/00_pileups.table".format(tumor,normal)],[gatk4,tumor,exomeRegions,gnomadVCF,normal],getPileupSummariesCMD)
calculateContaminationCMD = "${SOURCES[0]} CalculateContamination -I ${SOURCES[1]} -O $TARGET"
calculateContamination = env.Command(["Variants/Mutect2/{}_{}/00_contamination.table".format(tumor,normal)],[gatk4,getPileupSummaries],calculateContaminationCMD)
#######################################
##### Variant Calling with MuTect2 ####
#######################################
if "mutect2" in variantCallersList:
muTect2CMD = "${SOURCES[0]} Mutect2 -R ${SOURCES[1]} -I ${SOURCES[2]}/07_recalibrated.bam -tumor ${SOURCES[2]} -I ${SOURCES[3]}/07_recalibrated.bam -normal ${SOURCES[3]} -L ${SOURCES[4]} -O $TARGET --germline-resource ${SOURCES[5]}" + " {}".format(mutect2Parameters)+" -bamout Variants/Mutect2/${SOURCES[2]}_${SOURCES[3]}/01_mutect2.bam"+" --native-pair-hmm-threads {}".format(processors)
muTect2 = env.Command(["Variants/Mutect2/{}_{}/01_mutect2.vcf".format(tumor,normal)],[gatk4,reference,tumor,normal,exomeRegions,gnomadVCF],muTect2CMD)
if (contamination == "y" or contamination == "yes"):
filterMutectCallsCMD = "${SOURCES[0]} FilterMutectCalls -V ${SOURCES[1]} -contamination-table ${SOURCES[2]} -O $TARGET"
filterMutectCalls = env.Command(["Variants/Mutect2/{}_{}/02_mutect2_filtered.vcf".format(tumor,normal)],[gatk4,muTect2,calculateContamination],filterMutectCallsCMD)
else:
filterMutectCallsCMD = "${SOURCES[0]} FilterMutectCalls -V ${SOURCES[1]} -O $TARGET"
filterMutectCalls = env.Command(["Variants/Mutect2/{}_{}/02_mutect2_filtered.vcf".format(tumor,normal)],[gatk4,muTect2],filterMutectCallsCMD)
#Annotation by snpEff
mutect2EFFCMD = "java -jar /tmp/snpEff/snpEff.jar eff -v {} -stats Variants/Mutect2/{}_{}/snpEff_summary.html -dataDir {} $SOURCE > $TARGET".format(snpeffGenomeVersion,tumor,normal,snpeffDir)
mutect2EFF = env.Command(["Variants/Mutect2/{}_{}/03_mutect2_snpeff.vcf".format(tumor,normal),
"Variants/Mutect2/{}_{}/snpEff_summary.genes.txt".format(tumor,normal),
"Variants/Mutect2/{}_{}/snpEff_summary.html".format(tumor,normal)],[filterMutectCalls],mutect2EFFCMD)
# Annotation with ClinVar
mutect2ClinvarCMD = "java -Xmx2g -jar /tmp/snpEff/SnpSift.jar annotate -info CLNSIG,CLNDN {} $SOURCE > $TARGET".format(clinvar)
mutect2Clinvar = env.Command(["Variants/Mutect2/{}_{}/04_mutect2_clinvar.vcf".format(tumor,normal)], [mutect2EFF], mutect2ClinvarCMD)
# Cosmic coding variants annotation
mutect2CosmicCMD = "java -Xmx4g -jar /tmp/snpEff/SnpSift.jar annotate {} $SOURCE > $TARGET".format(cosmic)
mutect2Cosmic = env.Command(["Variants/Mutect2/{}_{}/05_mutect2_cosmic.vcf".format(tumor,normal)], [mutect2Clinvar], mutect2CosmicCMD)
# dbSNP annotation
mutect2DbsnpCMD = "java -Xmx4g -jar /tmp/snpEff/SnpSift.jar annotate {} $SOURCE > $TARGET".format(dbsnpVCF)
mutect2Dbsnp = env.Command(["Variants/Mutect2/{}_{}/06_mutect2_dbsnp.vcf".format(tumor,normal)], [mutect2Cosmic], mutect2DbsnpCMD)
# gnomAD annotation
mutect2gnomadCMD = "java -Xmx4g -jar /tmp/snpEff/SnpSift.jar annotate {} $SOURCE > $TARGET".format(gnomadVCF)
mutect2gnomad = env.Command(["Variants/Mutect2/{}_{}/07_mutect2_gnomad.vcf".format(tumor,normal)], [mutect2Dbsnp], mutect2gnomadCMD)
# Filtering variants
mutect2FilteringCMD = "python /tmp/mutect-vcf-selector.py -f $SOURCE -c /annotations/cancer_gene_census.csv -e > $TARGET"
mutect2Filtering = env.Command(["Variants/Mutect2/{}_{}/08_mutect2_filtered.vcf".format(tumor,normal)], [mutect2gnomad], mutect2FilteringCMD)
# Tag variant type
mutect2VariantTypeCMD= "java -Xmx4g -jar /tmp/snpEff/SnpSift.jar varType $SOURCE > $TARGET"
mutect2VariantType = env.Command(["Variants/Mutect2/{}_{}/08.5_mutect2_vartype.vcf".format(tumor,normal)], [mutect2Filtering], mutect2VariantTypeCMD)
# Selection of snps
mutect2snpCMD = "java -Xmx4g -jar /tmp/snpEff/SnpSift.jar filter \"(VARTYPE has 'SNP') | (VARTYPE has 'MNP')\" $SOURCE > $TARGET"
mutect2snp = env.Command(["Variants/Mutect2/{}_{}/09_mutect2_snps.vcf".format(tumor,normal)], [mutect2VariantType], mutect2snpCMD)
# Selection of indels
mutect2indelsCMD = "java -jar /tmp/snpEff/SnpSift.jar filter \"(VARTYPE has 'INS') | (VARTYPE has 'DEL') | (VARTYPE has 'MIXED')\" $SOURCE > $TARGET"
mutect2indels = env.Command(["Variants/Mutect2/{}_{}/09_mutect2_indels.vcf".format(tumor,normal)], [mutect2VariantType], mutect2indelsCMD)
# Copy latest created .vcf of snps to VCF directory
copyMutect2AnnotatedSnpsCMD = "cp $SOURCE $TARGET"
copyMutect2AnnotatedSnps = env.Command(["VCF/{}_{}_mutect2_snps.vcf".format(tumor,normal)], [mutect2snp], copyMutect2AnnotatedSnpsCMD)
# Copy latest created .vcf of indels to VCF directory
copyMutect2AnnotatedIndelsCMD = "cp $SOURCE $TARGET"
copyMutect2AnnotatedIndels = env.Command(["VCF/{}_{}_mutect2_indels.vcf".format(tumor,normal)], [mutect2indels], copyMutect2AnnotatedIndelsCMD)
#######################################
#### Variant Calling with Strelka2 ####
#######################################
if "strelka2" in variantCallersList:
strelka2configurationCMD = "python /tmp/strelka/bin/configureStrelkaSomaticWorkflow.py --normalBam ${SOURCES[0]}/07_recalibrated.bam --tumorBam ${SOURCES[1]}/07_recalibrated.bam --referenceFasta ${SOURCES[2]} --runDir $TARGET --callRegions ${SOURCES[3]}.gz --exome"
strelka2configuration = env.Command(["Variants/Strelka2/{}_{}/strelkaWorkflow".format(tumor,normal)],[normal,tumor,reference,exomeRegions],strelka2configurationCMD)
strelka2runCMD = "python $SOURCE/runWorkflow.py -m local"+" -j {}".format(processors)+" > $TARGET"
strelka2run = env.Command(["Variants/Strelka2/{}_{}/00_strelka2.done.txt".format(tumor,normal)],[strelka2configuration],strelka2runCMD)
selectVariantsStrelka2snvsCMD = "${SOURCES[0]}"+" SelectVariants -R ${SOURCES[1]}"+" -V Variants/Strelka2/{}_{}/strelkaWorkflow/results/variants/somatic.snvs.vcf.gz".format(tumor,normal)+" -O $TARGET --exclude-filtered"
selectVariantsStrelka2snvs = env.Command(["Variants/Strelka2/{}_{}/01_strelka2_snvs_pass.vcf".format(tumor,normal)],[gatk4,reference,strelka2run],selectVariantsStrelka2snvsCMD)
selectVariantsStrelka2indelsCMD = "${SOURCES[0]}"+" SelectVariants -R ${SOURCES[1]}"+" -V Variants/Strelka2/{}_{}/strelkaWorkflow/results/variants/somatic.indels.vcf.gz".format(tumor,normal)+" -O $TARGET --exclude-filtered"
selectVariantsStrelka2indels = env.Command(["Variants/Strelka2/{}_{}/02_strelka2_indels_pass.vcf".format(tumor,normal)],[gatk4,reference,selectVariantsStrelka2snvs],selectVariantsStrelka2indelsCMD)
#Annotation by snpEff
strelka2EFFCMD = "java -jar /tmp/snpEff/snpEff.jar eff -v {} -stats Variants/Strelka2/{}_{}/snpEff_summary.html -dataDir {} $SOURCE > $TARGET".format(snpeffGenomeVersion,tumor,normal,snpeffDir)
strelka2EFF = env.Command(["Variants/Strelka2/{}_{}/03_strelka2_snvs_snpeff.vcf".format(tumor,normal),
"Variants/Strelka2/{}_{}/snpEff_summary.genes.txt".format(tumor,normal),
"Variants/Strelka2/{}_{}/snpEff_summary.html".format(tumor,normal)],[selectVariantsStrelka2snvs], strelka2EFFCMD )
strelka2indelsEFFCMD = "java -jar /tmp/snpEff/snpEff.jar eff -v {} -stats Variants/Strelka2/{}_{}/snpEff_indels_summary.html -dataDir {} $SOURCE > $TARGET".format(snpeffGenomeVersion,tumor,normal,snpeffDir)
strelka2indelsEFF = env.Command(["Variants/Strelka2/{}_{}/03_strelka2_indels_snpeff.vcf".format(tumor,normal),
"Variants/Strelka2/{}_{}/snpEff_indels_summary.genes.txt".format(tumor,normal),
"Variants/Strelka2/{}_{}/snpEff_indels_summary.html".format(tumor,normal)],[selectVariantsStrelka2indels], strelka2indelsEFFCMD )
# Annotation with ClinVar
strelka2clinvarCmd = "java -Xmx4g -jar /tmp/snpEff/SnpSift.jar annotate -info CLNSIG,CLNDN {} $SOURCE > $TARGET".format(clinvar)
strelka2clinvar = env.Command(["Variants/Strelka2/{}_{}/04_strelka2_snvs_clinvar.vcf".format(tumor,normal)], [strelka2EFF], strelka2clinvarCmd)
strelka2indelsclinvarCmd = "java -Xmx4g -jar /tmp/snpEff/SnpSift.jar annotate -info CLNSIG,CLNDN {} $SOURCE > $TARGET".format(clinvar)
strelka2indelsclinvar = env.Command(["Variants/Strelka2/{}_{}/04_strelka2_indels_clinvar.vcf".format(tumor,normal)], [strelka2indelsEFF], strelka2indelsclinvarCmd)
# Cosmic coding variants annotation
cosmicCodingCmd = "java -Xmx4g -jar /tmp/snpEff/SnpSift.jar annotate {} $SOURCE > $TARGET".format(cosmic)
cosmicCoding = env.Command(["Variants/Strelka2/{}_{}/05_strelka2_snvs_cosmic.vcf".format(tumor,normal)], [strelka2EFF], cosmicCodingCmd)
cosmicindelsCodingCmd = "java -Xmx4g -jar /tmp/snpEff/SnpSift.jar annotate {} $SOURCE > $TARGET".format(cosmic)
cosmicindelsCoding = env.Command(["Variants/Strelka2/{}_{}/05_strelka2_indels_cosmic.vcf".format(tumor,normal)], [strelka2indelsclinvar], cosmicindelsCodingCmd)
# dbSNP annotation
strelka2DbsnpCMD = "java -Xmx4g -jar /tmp/snpEff/SnpSift.jar annotate {} $SOURCE > $TARGET".format(dbsnpVCF)
strelka2Dbsnp = env.Command(["Variants/Strelka2/{}_{}/06_strelka2_snvs_dbsnp.vcf".format(tumor,normal)], [cosmicCoding], strelka2DbsnpCMD)
strelka2indelsDbsnpCMD = "java -Xmx4g -jar /tmp/snpEff/SnpSift.jar annotate {} $SOURCE > $TARGET".format(dbsnpVCF)
strelka2indelsDbsnp = env.Command(["Variants/Strelka2/{}_{}/06_strelka2_indels_dbsnp.vcf".format(tumor,normal)], [cosmicindelsCoding], strelka2indelsDbsnpCMD)
# snps gnomAD annotation
strelka2gnomadCMD = "java -Xmx4g -jar /tmp/snpEff/SnpSift.jar annotate {} $SOURCE > $TARGET".format(gnomadVCF)
strelka2gnomad = env.Command(["Variants/Strelka2/{}_{}/07_strelka2_snvs_gnomad.vcf".format(tumor,normal)], [strelka2Dbsnp], strelka2gnomadCMD)
strelka2GenotypeConversionCMD = "update_GT_Strelka2.py -v $SOURCE -t snps > $TARGET"
strelka2GenotypeConversion = env.Command(["Variants/Strelka2/{}_{}/08_strelka2_snvs_gt.vcf".format(tumor,normal)], [strelka2gnomad], strelka2GenotypeConversionCMD)
# Indels gnomad annotation
strelka2indelsgnomadCMD = "java -Xmx4g -jar /tmp/snpEff/SnpSift.jar annotate {} $SOURCE > $TARGET".format(gnomadVCF)
strelka2indelsgnomad = env.Command(["Variants/Strelka2/{}_{}/07_strelka2_indels_gnomad.vcf".format(tumor,normal)], [strelka2indelsDbsnp], strelka2indelsgnomadCMD)
strelka2GenotypeConversionIndelsCMD = "update_GT_Strelka2.py -v $SOURCE -t indels > $TARGET"
strelka2GenotypeConversionIndels = env.Command(["Variants/Strelka2/{}_{}/08_strelka2_indels_gt.vcf".format(tumor,normal)], [strelka2indelsgnomad], strelka2GenotypeConversionIndelsCMD)
# Move latest created vcf of snps to VCF directory
copyStrelka2SnpsAnnotatedCMD = "cp $SOURCE $TARGET"
copyStrelka2SnpsAnnotated = env.Command(["VCF/{}_{}_strelka2_snps.vcf".format(tumor,normal)], [strelka2GenotypeConversion], copyStrelka2SnpsAnnotatedCMD)
# Move latest created vcf of indels to VCF directory
copyStrelka2IndelsAnnotatedCMD = "cp $SOURCE $TARGET"
copyStrelka2IndelsAnnotated = env.Command(["VCF/{}_{}_strelka2_indels.vcf".format(tumor,normal)], [strelka2GenotypeConversionIndels], copyStrelka2IndelsAnnotatedCMD)
######################################
#### Variant calling with VarScan ####
######################################
if "varscan" in variantCallersList:
varscanCMD = 'java -jar /tmp/VarScan.jar somatic ' \
' <(samtools mpileup -f ${SOURCES[0]} /working/${SOURCES[1]}/07_recalibrated.bam) ' \
' <(samtools mpileup -f ${SOURCES[0]} /working/${SOURCES[2]}/07_recalibrated.bam) ' + \
' Variants/VarScan/{}_{}/01_varscan ' \
' --min-coverage 10 ' \
' --somatic-p-value 0.05 ' \
' --p-value 0.99 ' \
' --min-var-freq 0.02 {}'.format(tumor,normal,varScanParameters)
varscan = env.Command(["Variants/VarScan/{}_{}/01_varscan.indel".format(tumor,normal), \
"Variants/VarScan/{}_{}/01_varscan.snp".format(tumor,normal)], \
[reference,normal,tumor], \
varscanCMD)
filterVarScanSnpsCMD = 'java -jar /tmp/VarScan.jar somaticFilter ' \
'Variants/VarScan/{}_{}/01_varscan.snp ' \
' --min-coverage {} ' \
' --pvalue {} ' \
' --min-var-freq {} ' \
' --output-file Variants/VarScan/{}_{}/02_snp ' \
' && cp Variants/VarScan/{}_{}/02_snp '.format(tumor,normal,varScanMinCoverage,varScanPvalue,varScanMinVarFreq,tumor,normal,tumor,normal) + \
'${TARGET}' + ' && rm Variants/VarScan/{}_{}/02_snp'.format(tumor,normal)
filterVarScanSnps = env.Command(["Variants/VarScan/{}_{}/02_varscan_snp_filtered.snp".format(tumor,normal)], \
[reference,normal,tumor,varscan], \
filterVarScanSnpsCMD)
filterVarScanIndelCMD = 'java -jar /tmp/VarScan.jar somaticFilter ' \
'Variants/VarScan/{}_{}/01_varscan.indel ' \
' --min-coverage {} ' \
' --pvalue {} ' \
' --min-var-freq {} ' \
' --output-file Variants/VarScan/{}_{}/02_indel && cp Variants/VarScan/{}_{}/02_indel '.format(tumor,normal,varScanMinCoverage,varScanPvalue,varScanMinVarFreq,tumor,normal,tumor,normal) + \
'${TARGET}' + \
' && rm Variants/VarScan/{}_{}/02_indel'.format(tumor,normal)
filterVarScanIndel = env.Command(["Variants/VarScan/{}_{}/02_varscan_indel_filtered.snp".format(tumor,normal)], [reference,normal,tumor,varscan], filterVarScanIndelCMD)
snpsToVCFCMD = 'python /tmp/vs_format_converter.py ' \
'Variants/VarScan/{}_{}/02_varscan_snp_filtered.snp > $TARGET'.format(tumor,normal)
snpsToVCF = env.Command(["Variants/VarScan/{}_{}/03_varscan_snp_filtered.vcf".format(tumor,normal)], [filterVarScanSnps], snpsToVCFCMD)
snpsToIndelsCMD = 'python /tmp/vs_format_converter.py ' \
'Variants/VarScan/{}_{}/02_varscan_indel_filtered.snp ' \
'> $TARGET'.format(tumor,normal)
snpsToIndels = env.Command(["Variants/VarScan/{}_{}/03_varscan_indels_filtered.vcf".format(tumor,normal)], [filterVarScanIndel], snpsToIndelsCMD)
#Annotation by snpEff
varscansnpEFFCMD = "java -jar /tmp/snpEff/snpEff.jar eff -v {} -stats Variants/VarScan/{}_{}/snpEff_summary.html -dataDir {} $SOURCE > $TARGET".format(snpeffGenomeVersion,tumor,normal,snpeffDir)
varscansnpEFF = env.Command(["Variants/VarScan/{}_{}/04_varscan_snp_snpeff.vcf".format(tumor,normal),
"Variants/VarScan/{}_{}/snpEff_summary.genes.txt".format(tumor,normal),
"Variants/VarScan/{}_{}/snpEff_summary.html".format(tumor,normal)],[snpsToVCF],varscansnpEFFCMD)
varscanindelsEFFCMD = "java -jar /tmp/snpEff/snpEff.jar eff -v {} -stats Variants/VarScan/{}_{}/snpEff_indels_summary.html -dataDir {} $SOURCE > $TARGET".format(snpeffGenomeVersion,tumor,normal,snpeffDir)
varscanindelsEFF = env.Command(["Variants/VarScan/{}_{}/04_varscan_indels_snpeff.vcf".format(tumor,normal),
"Variants/VarScan/{}_{}/snpEff_indels_summary.genes.txt".format(tumor,normal),
"Variants/VarScan/{}_{}/snpEff_indels_summary.html".format(tumor,normal)],[snpsToIndels],varscanindelsEFFCMD)
# Annotation with ClinVar
varscanclinvarCMD = "java -Xmx2g -jar /tmp/snpEff/SnpSift.jar annotate -info CLNSIG,CLNDN {} $SOURCE > $TARGET".format(clinvar)
varscanclinvar = env.Command(["Variants/VarScan/{}_{}/05_varscan_snp_clinvar.vcf".format(tumor,normal)], [varscansnpEFF], varscanclinvarCMD)
varscanindelsclinvarCMD = "java -Xmx2g -jar /tmp/snpEff/SnpSift.jar annotate -info CLNSIG,CLNDN {} $SOURCE > $TARGET".format(clinvar)
varscanindelsclinvar = env.Command(["Variants/VarScan/{}_{}/05_varscan_indels_clinvar.vcf".format(tumor,normal)], [varscanindelsEFF], varscanindelsclinvarCMD)
# Cosmic coding variants annotation
varscanCosmicCodingCMD = "java -Xmx4g -jar /tmp/snpEff/SnpSift.jar annotate {} $SOURCE > $TARGET".format(cosmic)
varscanCosmicCoding = env.Command(["Variants/VarScan/{}_{}/06_varscan_snp_cosmic.vcf".format(tumor,normal)], [varscanclinvar], varscanCosmicCodingCMD)
varscanindelsCosmicCodingCMD = "java -Xmx4g -jar /tmp/snpEff/SnpSift.jar annotate {} $SOURCE > $TARGET".format(cosmic)
varscanindelsCosmicCoding = env.Command(["Variants/VarScan/{}_{}/06_varscan_indels_cosmic.vcf".format(tumor,normal)], [varscanindelsclinvar], varscanindelsCosmicCodingCMD)
# dbSNP annotation
varscanDbsnpCMD = "java -Xmx4g -jar /tmp/snpEff/SnpSift.jar annotate {} $SOURCE > $TARGET".format(dbsnpVCF)
varscanDbsnp = env.Command(["Variants/VarScan/{}_{}/07_varscan_snp_dbsnp.vcf".format(tumor,normal)], [varscanCosmicCoding], varscanDbsnpCMD)
varscanindelsDbsnpCMD = "java -Xmx4g -jar /tmp/snpEff/SnpSift.jar annotate {} $SOURCE > $TARGET".format(dbsnpVCF)
varscanindelsDbsnp = env.Command(["Variants/VarScan/{}_{}/07_varscan_indels_dbsnp.vcf".format(tumor,normal)], [varscanindelsCosmicCoding], varscanindelsDbsnpCMD)
# gnomAD annotation
varscangnomadCMD = "java -Xmx4g -jar /tmp/snpEff/SnpSift.jar annotate {} $SOURCE > $TARGET".format(gnomadVCF)
varscangnomad = env.Command(["Variants/VarScan/{}_{}/08_varscan_snp_gnomad.vcf".format(tumor,normal)], [varscanDbsnp], varscangnomadCMD)
varscanindelsgnomadCMD = "java -Xmx4g -jar /tmp/snpEff/SnpSift.jar annotate {} $SOURCE > $TARGET".format(gnomadVCF)
varscanindelsgnomad = env.Command(["Variants/VarScan/{}_{}/08_varscan_indels_gnomad.vcf".format(tumor,normal)], [varscanindelsDbsnp], varscanindelsgnomadCMD)
# Move latest created vcf of snps to VCF directory
copyVarscanAnnotatedCMD = "cp $SOURCE $TARGET"
copyVarscanAnnotated = env.Command(["VCF/{}_{}_varscan_snps.vcf".format(tumor,normal)], [varscangnomad], copyVarscanAnnotatedCMD)
# Move latest created vcf of indels to VCF directory
copyVarscanAnnotatedIndelsCMD = "cp $SOURCE $TARGET"
copyVarscanAnnotatedIndels = env.Command(["VCF/{}_{}_varscan_indels.vcf".format(tumor,normal)], [varscanindelsgnomad], copyVarscanAnnotatedIndelsCMD)
# ===========================================================================================
variantCallersString = "java -Xmx4g -jar /tmp/gatk3/GenomeAnalysisTK.jar -T CombineVariants -R {} ".format(reference)
variantCallersIndelsString = variantCallersString
lastStepOfVariantCallerList = []
lastStepOfIndelCallerList = []
lastStepOfVariantCaller = {}
lastStepOfIndelVariantCaller = {}
for variantCaller in variantCallersList:
if variantCaller == "varscan":
lastStepOfVariantCaller["varscan"] = copyVarscanAnnotated[0]
lastStepOfIndelVariantCaller["varscan"] = copyVarscanAnnotatedIndels[0]
if variantCaller == "strelka2":
lastStepOfVariantCaller["strelka2"] = copyStrelka2SnpsAnnotated[0]
lastStepOfIndelVariantCaller["strelka2"] = copyStrelka2IndelsAnnotated[0]
if variantCaller == "mutect2":
lastStepOfVariantCaller["mutect2"] = copyMutect2AnnotatedSnps[0]
lastStepOfIndelVariantCaller["mutect2"] = copyMutect2AnnotatedIndels[0]
for variantCaller in variantCallersList:
variantCallersString = variantCallersString + " --variant:{} /working/VCF/{}_{}_{}_snps.vcf ".format(variantCaller,tumor,normal,variantCaller)
lastStepOfVariantCallerList.append(lastStepOfVariantCaller.get(variantCaller))
variantCallersIndelsString = variantCallersIndelsString + " --variant:{} /working/VCF/{}_{}_{}_indels.vcf ".format(variantCaller,tumor,normal,variantCaller)
lastStepOfIndelCallerList.append(lastStepOfIndelVariantCaller.get(variantCaller))
variantCallersList.sort()
lastStepOfIndelCallerList.sort()
variantCallerListIndels = variantCallersList[0:]
variantCallersStringCMD = variantCallersString + " -o VCF/Combined_VCFs_by_sample/{}_{}_merged_snps.vcf -genotypeMergeOptions PRIORITIZE -priority {}".format(tumor,normal, ",".join(variantCallersList))
combineVCFsSnps = env.Command(["VCF/Combined_VCFs_by_sample/{}_{}_merged_snps.vcf".format(tumor,normal)], lastStepOfVariantCallerList, variantCallersStringCMD)
variantCallersIndelsString = variantCallersIndelsString + " -o VCF/Combined_VCFs_by_sample/{}_{}_merged_indels.vcf -genotypeMergeOptions PRIORITIZE -priority {}".format(tumor,normal,",".join(variantCallerListIndels))
combineVCFsIndels = env.Command(["VCF/Combined_VCFs_by_sample/{}_{}_merged_indels.vcf".format(tumor,normal)], lastStepOfIndelCallerList, variantCallersIndelsString )