Empty vcf when generating PON with MuTect2

HoxusHoxus Member
edited April 2017 in Ask the GATK team

Hi,

First, sorry if it's a duplicate question. I have looked for solution, but didn't find.

I have 8 exomes data from 4 patient: normal and tumor.
I ran MuTect2 (in GTAK 3.7) in normal/tumor mode, and it works well.

But now, I would like to generate a Panel of Normal.
Here the log:

INFO  16:48:48,998 HelpFormatter - -------------------------------------------------------------------------------- 
INFO  16:48:49,000 HelpFormatter - The Genome Analysis Toolkit (GATK) v3.7-0-gcfedb67, Compiled 2016/12/12 11:21:18 
INFO  16:48:49,001 HelpFormatter - Copyright (c) 2010-2016 The Broad Institute 
INFO  16:48:49,001 HelpFormatter - For support and documentation go to https://software.broadinstitute.org/gatk 
INFO  16:48:49,001 HelpFormatter - [Mon Apr 03 16:48:48 CEST 2017] Executing on Linux 4.4.0-62-generic amd64 
INFO  16:48:49,001 HelpFormatter - Java HotSpot(TM) 64-Bit Server VM 1.8.0_121-b13 
INFO  16:48:49,004 HelpFormatter - Program Args: -T MuTect2 -nct 2 -R /media/raid/genomic_DB//hg19/hg19.fasta -I:normal bam/WES_8N.sorted.dedup.recalib.bam -L /media/raid/genomic_DB//hg19/bed/hg19_Gene_table.bed --dbsnp /media/raid/genomic_DB//hg19/GATK_bundle/dbsnp_138.hg19.vcf --cosmic /media/raid/genomic_DB//hg19/GATK_bundle/Cosmic.hg19.vcf --artifact_detection_mode -o var/WES_8_N.vcf 
INFO  16:48:49,007 HelpFormatter - Executing as [email protected] on Linux 4.4.0-62-generic amd64; Java HotSpot(TM) 64-Bit Server VM 1.8.0_121-b13. 
INFO  16:48:49,007 HelpFormatter - Date/Time: 2017/04/03 16:48:48 
INFO  16:48:49,008 HelpFormatter - -------------------------------------------------------------------------------- 
INFO  16:48:49,008 HelpFormatter - -------------------------------------------------------------------------------- 
INFO  16:48:49,023 GenomeAnalysisEngine - Strictness is SILENT 
INFO  16:48:49,133 GenomeAnalysisEngine - Downsampling Settings: Method: BY_SAMPLE, Target Coverage: 1000 
INFO  16:48:49,140 SAMDataSource$SAMReaders - Initializing SAMRecords in serial 
INFO  16:48:49,160 SAMDataSource$SAMReaders - Done initializing BAM readers: total time 0.02 
INFO  16:48:50,273 IntervalUtils - Processing 1325431593 bp from intervals 
INFO  16:48:50,285 MicroScheduler - Running the GATK in parallel mode with 2 total threads, 2 CPU thread(s) for each of 1 data thread(s), of 56 processors available on this machine 
INFO  16:48:50,340 GenomeAnalysisEngine - Preparing for traversal over 1 BAM files 
INFO  16:48:50,465 GenomeAnalysisEngine - Done preparing for traversal 
INFO  16:48:50,466 ProgressMeter - [INITIALIZATION COMPLETE; STARTING PROCESSING] 
INFO  16:48:50,466 ProgressMeter -                 |      processed |    time |         per 1M |           |   total | remaining 
INFO  16:48:50,466 ProgressMeter -        Location | active regions | elapsed | active regions | completed | runtime |   runtime 
INFO  16:48:50,532 MuTect2 - Using global mismapping rate of 45 => -4.5 in log10 likelihood units 
INFO  16:48:50,533 PairHMM - Performance profiling for PairHMM is disabled because the program is being run with multiple threads (-nct>1) option
Profiling is enabled only when running in single thread mode

INFO  16:49:20,470 ProgressMeter -   chr1:14958958     5.97786178E8    30.0 s            0.0 s        0.6%    82.1 m      81.6 m 
INFO  16:49:50,473 ProgressMeter -   chr1:26627596    2.516091919E9    60.0 s            0.0 s        1.1%    88.4 m      87.4 m 
INFO  16:50:20,474 ProgressMeter -   chr1:42766671    6.139828701E9    90.0 s            0.0 s        1.7%    85.9 m      84.4 m 
INFO  16:50:50,476 ProgressMeter -   chr1:62963628   1.0743175026E10   120.0 s            0.0 s        2.7%    73.3 m      71.3 m 
INFO  16:51:20,478 ProgressMeter -   chr1:91817153   1.5892962774E10     2.5 m            0.0 s        3.8%    66.3 m      63.8 m 
INFO  16:51:50,480 ProgressMeter -  chr1:117093107   2.5856520721E10     3.0 m            0.0 s        4.6%    65.4 m      62.4 m 
INFO  16:52:20,482 ProgressMeter -  chr1:156181686   4.2774755347E10     3.5 m            0.0 s        5.3%    65.5 m      62.0 m 
INFO  16:52:50,484 ProgressMeter -  chr1:178832743   5.9965019186E10     4.0 m            0.0 s        6.2%    64.3 m      60.3 m 
INFO  16:53:20,486 ProgressMeter -  chr1:205910763   7.6568008961E10     4.5 m            0.0 s        7.1%    63.5 m      59.0 m 
INFO  16:53:50,487 ProgressMeter -  chr1:231612271   9.5039378738E10     5.0 m            0.0 s        8.1%    62.0 m      57.0 m 
INFO  16:54:20,489 ProgressMeter -   chr2:10978103   1.08452450175E11     5.5 m            0.0 s        9.1%    60.2 m      54.7 m 
...
...
*MuTect2 running, without any warnings, so I delete lines to keep message shorter*
...
...
INFO  18:55:50,952 ProgressMeter -   chrX:76856153   6.28621417292E11     2.1 h            0.0 s       97.8%     2.2 h       2.8 m 
INFO  18:56:20,953 ProgressMeter -   chrX:76877471   6.28621417292E11     2.1 h            0.0 s       97.8%     2.2 h       2.9 m 
INFO  18:56:50,954 ProgressMeter -   chrX:76893542   6.28621417292E11     2.1 h            0.0 s       97.8%     2.2 h       2.9 m 
INFO  18:57:20,956 ProgressMeter -   chrX:76906228   6.28621417292E11     2.1 h            0.0 s       97.8%     2.2 h       2.9 m 
INFO  18:57:50,958 ProgressMeter -   chrX:76917364   6.28621417292E11     2.2 h            0.0 s       97.8%     2.2 h       2.9 m 
INFO  18:58:20,959 ProgressMeter -   chrX:76927855   6.28621417292E11     2.2 h            0.0 s       97.8%     2.2 h       2.9 m 
INFO  18:58:50,961 ProgressMeter -   chrX:76937585   6.28621417292E11     2.2 h            0.0 s       97.8%     2.2 h       2.9 m 
INFO  18:59:20,962 ProgressMeter -   chrX:76946451   6.28621417292E11     2.2 h            0.0 s       97.8%     2.2 h       2.9 m 
INFO  18:59:50,964 ProgressMeter -   chrX:76954953   6.28621417292E11     2.2 h            0.0 s       97.8%     2.2 h       2.9 m 
INFO  19:00:20,965 ProgressMeter -   chrX:76963009   6.28621417292E11     2.2 h            0.0 s       97.8%     2.2 h       2.9 m 
INFO  19:00:50,966 ProgressMeter -   chrX:76970682   6.28621417292E11     2.2 h            0.0 s       97.8%     2.2 h       2.9 m 
INFO  19:01:20,967 ProgressMeter -  chrX:118733701   6.34550108037E11     2.2 h            0.0 s       98.9%     2.2 h      92.0 s 
Using AVX accelerated implementation of PairHMM
INFO  19:01:44,249 VectorLoglessPairHMM - libVectorLoglessPairHMM unpacked successfully from GATK jar file 
INFO  19:01:44,249 VectorLoglessPairHMM - Using vectorized implementation of PairHMM 
INFO  19:01:44,250 MuTect2 - Ran local assembly on 0 active regions 
INFO  19:01:44,252 ProgressMeter -            done   6.48115744256E11     2.2 h            0.0 s      100.0%     2.2 h       0.0 s 
INFO  19:01:44,253 ProgressMeter - Total runtime 7973.79 secs, 132.90 min, 2.21 hours 
INFO  19:01:44,253 MicroScheduler - 386106 reads were filtered out during the traversal out of approximately 105750521 total reads (0.37%) 
INFO  19:01:44,253 MicroScheduler -   -> 0 reads (0.00% of total) failing BadCigarFilter 
INFO  19:01:44,253 MicroScheduler -   -> 371854 reads (0.35% of total) failing DuplicateReadFilter 
INFO  19:01:44,253 MicroScheduler -   -> 0 reads (0.00% of total) failing FailsVendorQualityCheckFilter 
INFO  19:01:44,254 MicroScheduler -   -> 0 reads (0.00% of total) failing MalformedReadFilter 
INFO  19:01:44,254 MicroScheduler -   -> 0 reads (0.00% of total) failing MappingQualityUnavailableFilter 
INFO  19:01:44,254 MicroScheduler -   -> 14252 reads (0.01% of total) failing NotPrimaryAlignmentFilter 
INFO  19:01:44,254 MicroScheduler -   -> 0 reads (0.00% of total) failing UnmappedReadFilter 
------------------------------------------------------------------------------------------
Done. There were no warn messages.
------------------------------------------------------------------------------------------

At the end, I have a vcf without any variant, only header appear.
I have looked for solutions, and it seems I have the same problem like here:
1. http://gatkforums.broadinstitute.org/gatk/discussion/6085/generating-pool-of-normal
2. https://gatkforums.broadinstitute.org/wdl/discussion/6690/mutect2-tumor-only-mode-empty-vcfs

As in previous post, I read the same strange info in my log:

INFO  19:01:44,250 MuTect2 - Ran local assembly on 0 active regions

So I have tried a nightly build, as recommended in previous post:

INFO  16:09:07,202 HelpFormatter - --------------------------------------------------------------------------------------------- 
INFO  16:09:07,204 HelpFormatter - The Genome Analysis Toolkit (GATK) vnightly-2017-04-04-g34bd8a3, Compiled 2017/04/04 00:01:14 
INFO  16:09:07,204 HelpFormatter - Copyright (c) 2010-2016 The Broad Institute 
INFO  16:09:07,204 HelpFormatter - For support and documentation go to https://software.broadinstitute.org/gatk 
INFO  16:09:07,204 HelpFormatter - [Tue Apr 04 16:09:07 CEST 2017] Executing on Linux 4.4.0-62-generic amd64 
INFO  16:09:07,204 HelpFormatter - Java HotSpot(TM) 64-Bit Server VM 1.8.0_121-b13 
INFO  16:09:07,208 HelpFormatter - Program Args: -T MuTect2 -nct 2 -R /media/raid/genomic_DB//hg19/hg19.fasta -I:normal bam/WES_26N.sorted.dedup.recalib.bam -L /media/raid/genomic_DB//hg19/bed/hg19_Gene_table.bed --dbsnp /media/raid/genomic_DB//hg19/GATK_bundle/dbsnp_138.hg19.vcf --cosmic /media/raid/genomic_DB//hg19/GATK_bundle/Cosmic.hg19.vcf --artifact_detection_mode -o var/WES_26_N.vcf 
INFO  16:09:07,210 HelpFormatter - Executing as [email protected] on Linux 4.4.0-62-generic amd64; Java HotSpot(TM) 64-Bit Server VM 1.8.0_121-b13. 
INFO  16:09:07,210 HelpFormatter - Date/Time: 2017/04/04 16:09:07 
INFO  16:09:07,210 HelpFormatter - --------------------------------------------------------------------------------------------- 
INFO  16:09:07,210 HelpFormatter - --------------------------------------------------------------------------------------------- 
INFO  16:09:07,225 GenomeAnalysisEngine - Strictness is SILENT 
INFO  16:09:07,340 GenomeAnalysisEngine - Downsampling Settings: Method: BY_SAMPLE, Target Coverage: 1000 
INFO  16:09:07,346 SAMDataSource$SAMReaders - Initializing SAMRecords in serial 
INFO  16:09:07,366 SAMDataSource$SAMReaders - Done initializing BAM readers: total time 0.02 
INFO  16:09:08,334 IntervalUtils - Processing 1325431593 bp from intervals 
INFO  16:09:08,346 MicroScheduler - Running the GATK in parallel mode with 2 total threads, 2 CPU thread(s) for each of 1 data thread(s), of 56 processors available on this machine 
INFO  16:09:08,399 GenomeAnalysisEngine - Preparing for traversal over 1 BAM files 
INFO  16:09:08,539 GenomeAnalysisEngine - Done preparing for traversal 
INFO  16:09:08,540 ProgressMeter - [INITIALIZATION COMPLETE; STARTING PROCESSING] 
INFO  16:09:08,540 ProgressMeter -                 |      processed |    time |         per 1M |           |   total | remaining 
INFO  16:09:08,542 ProgressMeter -        Location | active regions | elapsed | active regions | completed | runtime |   runtime 
INFO  16:09:08,610 MuTect2 - Using global mismapping rate of 45 => -4.5 in log10 likelihood units 
INFO  16:09:08,611 PairHMM - Performance profiling for PairHMM is disabled because the program is being run with multiple threads (-nct>1) option
Profiling is enabled only when running in single thread mode

INFO  16:09:38,546 ProgressMeter -   chr1:16563659     7.97486694E8    30.0 s            0.0 s        0.7%    71.0 m      70.5 m 
INFO  16:10:08,547 ProgressMeter -   chr1:36384738    4.675477702E9    60.0 s            0.0 s        1.5%    66.7 m      65.7 m 
INFO  16:10:38,549 ProgressMeter -   chr1:65120499   1.1110499512E10    90.0 s            0.0 s        2.8%    53.0 m      51.5 m 
INFO  16:11:08,551 ProgressMeter -  chr1:108509208   1.9618437838E10   120.0 s            0.0 s        4.3%    46.9 m      44.9 m 
...
...
again, I delete some lines without warnings to keep shorter
...
...
INFO  18:07:38,969 ProgressMeter -   chrX:76944672   6.28621417292E11   118.5 m            0.0 s       97.8%     2.0 h       2.7 m 
INFO  18:08:08,970 ProgressMeter -   chrX:76953314   6.28621417292E11   119.0 m            0.0 s       97.8%     2.0 h       2.7 m 
INFO  18:08:38,971 ProgressMeter -   chrX:76961411   6.28621417292E11   119.5 m            0.0 s       97.8%     2.0 h       2.7 m 
INFO  18:09:08,973 ProgressMeter -   chrX:76968843   6.28621417292E11   120.0 m            0.0 s       97.8%     2.0 h       2.7 m 
INFO  18:09:38,974 ProgressMeter -  chrX:113099932   6.33515778237E11     2.0 h            0.0 s       98.7%     2.0 h      93.0 s 
Using AVX accelerated implementation of PairHMM
INFO  18:10:00,807 VectorLoglessPairHMM - libVectorLoglessPairHMM unpacked successfully from GATK jar file 
INFO  18:10:00,808 VectorLoglessPairHMM - Using vectorized implementation of PairHMM 
INFO  18:10:00,809 MuTect2 - Ran local assembly on 0 active regions 
INFO  18:10:00,812 ProgressMeter -            done   6.48115744256E11     2.0 h            0.0 s      100.0%     2.0 h       0.0 s 
INFO  18:10:00,812 ProgressMeter - Total runtime 7252.27 secs, 120.87 min, 2.01 hours 
INFO  18:10:00,813 MicroScheduler - 231969 reads were filtered out during the traversal out of approximately 82616070 total reads (0.28%) 
INFO  18:10:00,813 MicroScheduler -   -> 0 reads (0.00% of total) failing BadCigarFilter 
INFO  18:10:00,813 MicroScheduler -   -> 215232 reads (0.26% of total) failing DuplicateReadFilter 
INFO  18:10:00,813 MicroScheduler -   -> 0 reads (0.00% of total) failing FailsVendorQualityCheckFilter 
INFO  18:10:00,813 MicroScheduler -   -> 0 reads (0.00% of total) failing MalformedReadFilter 
INFO  18:10:00,813 MicroScheduler -   -> 0 reads (0.00% of total) failing MappingQualityUnavailableFilter 
INFO  18:10:00,814 MicroScheduler -   -> 16737 reads (0.02% of total) failing NotPrimaryAlignmentFilter 
INFO  18:10:00,814 MicroScheduler -   -> 0 reads (0.00% of total) failing UnmappedReadFilter 
------------------------------------------------------------------------------------------
Done. There were no warn messages.
------------------------------------------------------------------------------------------

Nightly build doesn't work better.
Not shown here, but I have tried without -L myBedFile.bed, and also without -nct, and I get a empty vcf again.
I don't know what to do/try...

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