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hartwigmedical / hmftools

Various algorithms for analysing genomics data
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[FATAL] Error executing R script in Amber #505

Closed XinweiZhao202 closed 5 months ago

XinweiZhao202 commented 5 months ago

I'm sorry that I'm really struggling with this issue, therefore I have to bother. The problem occurs in the segmentation step, which needs to call Rscript, the following is the log from Amber 4.0, actually I tried Amber from version 3.1 to 4.0, in the earlier versions the error log is the /tmp folder and it shows: During startup - Warning messages: 1: Setting LC_MONETARY failed, using "C" 2: Setting LC_PAPER failed, using "C" 3: Setting LC_MEASUREMENT failed, using "C" Loading required package: BiocGenerics

Attaching package: ‘BiocGenerics’

The following objects are masked from ‘package:stats’:

IQR, mad, sd, var, xtabs

The following objects are masked from ‘package:base’:

anyDuplicated, aperm, append, as.data.frame, basename, cbind,
colnames, dirname, do.call, duplicated, eval, evalq, Filter, Find,
get, grep, grepl, intersect, is.unsorted, lapply, Map, mapply,
match, mget, order, paste, pmax, pmax.int, pmin, pmin.int,
Position, rank, rbind, Reduce, rownames, sapply, setdiff, sort,
table, tapply, union, unique, unsplit, which.max, which.min

Error: input in data column 2 (posistions) must be numeric Execution halted

the log from Amber 4.0 is as follows: java -jar ./bin/hmftools/amber_v4.0.rc1.jar -threads 24 -loci bin/hmftools/HMFtools-Resources_dna_pipeline_v5_33_38_hmf_dna_pipeline_resources.38_v5.33/copy_number/GermlineHetPon.38.vcf.gz -tumor IC009T -tumor_bam /on2/bowtie2_results/IC009T/IC009T.sort.bam -output_dir /on2/hmftools_results/amber_results -ref_genome Homo_sapiens.GRCh38.dna.primary_assembly.fa 18:32:25.344 [INFO ] Amber version 4.0 18:32:32.531 [INFO ] removed 0 blacklisted loci, 7249216 remaining 18:32:32.531 [INFO ] processing tumor germline heterozygous(7249216) and homozygous(0) sites 18:33:03.550 [INFO ] [## ] 11% complete 18:33:33.552 [INFO ] [### ] 17% complete 18:34:03.554 [INFO ] [##### ] 26% complete 18:34:33.555 [INFO ] [####### ] 35% complete 18:35:03.557 [INFO ] [######## ] 44% complete 18:35:33.558 [INFO ] [########## ] 52% complete 18:36:03.559 [INFO ] [############ ] 63% complete 18:36:33.560 [INFO ] [################ ] 80% complete 18:37:03.560 [INFO ] [################# ] 87% complete 18:37:33.561 [INFO ] [################## ] 94% complete 18:38:01.026 [INFO ] [####################] 100% complete 18:38:02.669 [INFO ] contamination sites median tumor depth(0) 18:38:02.672 [INFO ] no evidence of contamination 18:38:02.675 [INFO ] applying pcf segmentation During startup - Warning messages: 1: Setting LC_MONETARY failed, using "C" 2: Setting LC_PAPER failed, using "C" 3: Setting LC_MEASUREMENT failed, using "C"

Attaching package: ???dplyr???

The following objects are masked from ???package:stats???:

filter, lag

The following objects are masked from ???package:base???:

intersect, setdiff, setequal, union

Loading required package: BiocGenerics

Attaching package: ???BiocGenerics???

The following objects are masked from ???package:dplyr???:

combine, intersect, setdiff, union

The following objects are masked from ???package:stats???:

IQR, mad, sd, var, xtabs

The following objects are masked from ???package:base???:

anyDuplicated, aperm, append, as.data.frame, basename, cbind,
colnames, dirname, do.call, duplicated, eval, evalq, Filter, Find,
get, grep, grepl, intersect, is.unsorted, lapply, Map, mapply,
match, mget, order, paste, pmax, pmax.int, pmin, pmin.int,
Position, rank, rbind, Reduce, rownames, sapply, setdiff, sort,
table, tapply, union, unique, unsplit, which.max, which.min

Error: input in data column 2 (posistions) must be numeric Execution halted 18:38:04.486 [FATAL] Error executing R script. Exception in thread "main" java.io.IOException: R execution failed. Unable to complete segmentation. at com.hartwig.hmftools.amber.BAFSegmentation.applySegmentation(BAFSegmentation.java:23) at com.hartwig.hmftools.amber.ResultsWriter.persistBAF(ResultsWriter.java:43) at com.hartwig.hmftools.amber.AmberApplication.runTumorOnly(AmberApplication.java:220) at com.hartwig.hmftools.amber.AmberApplication.run(AmberApplication.java:72) at com.hartwig.hmftools.amber.AmberApplication.main(AmberApplication.java:300)

Thanks in advance and have a nice day!

Regards Xinwei

hongwingl commented 5 months ago

Would you be able to look at the IC009T.amber.baf.tsv.gz file and see if the position column is all numeric?

XinweiZhao202 commented 5 months ago

Dear Mr.Lee, thanks for your question, honestly in the IC009T.baf.tsv file no concrete content except the header, I copy the header for you and this is all my get from the file. The header is as follows: chromosome position tumorBAF tumorModifiedBAF tumorDepth normalBAF normalModifiedBAF normalDepth

Another report is maybe useful for you: COBALT V1.14 could run without error, but V1.15.1 and V1.15.2 shows an error "inconsistent chromosome name: 20 and chr20" bei V1.15.1,"inconsistent chromosome name: X and chrX" bei V1.15.2. Anyway, I will go on the job with COBALT V1.14. Thanks in Advance.

Regards Xinwei

XinweiZhao202 commented 5 months ago

I fix the locale setting of R and remove warnings and run AMBER V4.0 once again, but same error, and I also run V3.9 it shows same things as before, I'm not sure if the input is problmatic, and the sorted bam file from Bowtie2 could be used in other process, I'm confused about this. here is the log from V3.9: java -jar ./bin/hmftools/amber-3.9.jar -threads 24 -loci bin/hmftools/HMFtools-Resources_dna_pipeline_v5_33_38_hmf_dna_pipeline_resources.38_v5.33/copy_number/GermlineHetPon.38.vcf.gz -tumor IC009T -tumor_bam /on2/bowtie2_results/IC009T/IC009T.sort.bam -output_dir /on2/hmftools_results/amber_results -ref_genome_version V38 16:21:02.418 INFO [main] - AMBER version: 3.9 16:21:02.419 INFO [main] - Loading vcf file bin/hmftools/HMFtools-Resources_dna_pipeline_v5_33_38_hmf_dna_pipeline_resources.38_v5.33/copy_number/GermlineHetPon.38.vcf.gz 16:21:09.433 INFO [main] - loaded 7249216 baf loci 16:21:10.704 INFO [main] - removed 15285 blacklisted loci, 7233931 remaining 16:21:10.704 INFO [main] - Processing 7233931 germline heterozygous loci in tumor bam /on2/bowtie2_results/IC009T/IC009T.sort.bam 16:21:10.705 INFO [main] - Processing 0 germline homozygous loci in tumor bam /on2/bowtie2_results/IC009T/IC009T.sort.bam for contamination 16:21:12.637 INFO [main] - 7233931 loci, 144895 genome regions, min gap = 2000 Exception in thread "worker-0" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-1" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-2" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-3" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-4" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-5" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-6" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-7" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-8" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-9" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-10" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-11" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-12" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-13" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-14" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-15" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-16" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-17" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-18" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-19" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-20" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-21" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-22" java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) Exception in thread "worker-23" 16:21:12.794 INFO [main] - 24 bam reader threads started java.lang.IllegalArgumentException: Invalid reference index -1 at htsjdk.samtools.QueryInterval.(QueryInterval.java:24) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.query(SamReader.java:538) at htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter.queryOverlapping(SamReader.java:400) at com.hartwig.hmftools.amber.AsyncBamLociReader$BamReaderThread.run(AsyncBamLociReader.java:69) 16:21:12.802 INFO [main] - 24 bam reader threads finished 16:21:13.184 INFO [main] - Median tumor depth at potential contamination sites is 0 reads 16:21:13.188 INFO [main] - No evidence of contamination. 16:21:13.192 INFO [main] - Applying pcf segmentation 16:21:13.194 INFO [main] - Executing R script via command: Rscript /tmp/script2229724272457342144.R /on2/hmftools_results/amber_results/IC009T.amber.baf.tsv.gz /on2/hmftools_results/amber_results/IC009T.amber.baf.pcf 16:21:15.000 FATAL [main] - Error executing R script. Examine error file /tmp/bafSegmentation.R7374888497635738545.error for details. Exception in thread "main" java.io.IOException: R execution failed. Unable to complete segmentation. at com.hartwig.hmftools.amber.BAFSegmentation.applySegmentation(BAFSegmentation.java:23) at com.hartwig.hmftools.amber.AmberPersistence.persistBAF(AmberPersistence.java:46) at com.hartwig.hmftools.amber.AmberApplication.runTumorOnly(AmberApplication.java:143) at com.hartwig.hmftools.amber.AmberApplication.run(AmberApplication.java:71) at com.hartwig.hmftools.amber.AmberApplication.main(AmberApplication.java:238)

hongwingl commented 5 months ago

I see. The reason is that in HG38 chromosome name have chr prefix, chr1, chr2, chrX etc. However seems your bam file contig names do not have those. That is why the programs failed.

XinweiZhao202 commented 5 months ago

Thanks but in COBALT 1.14 runs fine, this could also cause the error in AMBER maybe? I would like to change from Bowtie2 to BWA, I will give you feedback afterwards.

Regards Xinwei

hongwingl commented 5 months ago

cobalt 1.14 probably did not run fine. It would have given empty output. Newer version of cobalt checked to make chromosome names were correct and thus output an error. HG38 we need chr prefix for chromosome, and no chr for HG37. Which reference genome did you use to align your bam?

XinweiZhao202 commented 5 months ago

the results from COBALT 1.14 are not empty, I check the IC009T.cobalt.ratio.tsv file and it looks strange, I attach the first 20 rows as follows, maybe they could not be used for next stage. Screenshot at 2024-01-21 15-28-15

I use HG38 as conference, and at this moment I'm trying to use BWA bam as input, still with HG 38

XinweiZhao202 commented 5 months ago

Dear Mr. Lee, I tried the bam file from BWA but the errors are same as before, so alignment tools are not problem, is that necessary to change reference genome from V38 to V37?

Regards Xinwei

hongwingl commented 5 months ago

Would you be able to show me the output of samtools view -H <bam-file>?

XinweiZhao202 commented 5 months ago

Dear Mr. Lee, I change reference from V38 to V37, and the job by AMBER 3.9 and COBALT 1.15.2 could run without error, now I guess the reason is reference version, but I'm not sure about the detailed location of the error, by AMBER 4.0 it shows the Rscript error "Error: input in data column 2 (posistions) must be numeric", which maybe interesting for you. Anyway, my question is solved.

as you asked, I would like to show you headers of the bam, but I only have the bam with GRCh37, because I delete the other bam to save space, I also want to see if there are different to bam with GRCh38, so I will run this job and come back tomorrow.

Regards Xinwei

hongwingl commented 5 months ago

Hi, Great to hear that V37 works for you. The reason is that in the V38 reference genome that you used, the chromosomes do not have chr prefix. If you still want to use V38, the recommended way is to use the reference genome that Hartwig provides.

XinweiZhao202 commented 5 months ago

Dear Mr. Lee, I get the headers of each bam file, as far as I see, I didn't get the difference from the chr name, maybe you could find something, anyway, thanks for your help and I could move on.

Regards Xinwei

GRCh37 samtools view -H IC009T.sort.bam @HD VN:1.5 SO:coordinate @SQ SN:1 LN:249250621 @SQ SN:10 LN:135534747 @SQ SN:11 LN:135006516 @SQ SN:12 LN:133851895 @SQ SN:13 LN:115169878 @SQ SN:14 LN:107349540 @SQ SN:15 LN:102531392 @SQ SN:16 LN:90354753 @SQ SN:17 LN:81195210 @SQ SN:18 LN:78077248 @SQ SN:19 LN:59128983 @SQ SN:2 LN:243199373 @SQ SN:20 LN:63025520 @SQ SN:21 LN:48129895 @SQ SN:22 LN:51304566 @SQ SN:3 LN:198022430 @SQ SN:4 LN:191154276 @SQ SN:5 LN:180915260 @SQ SN:6 LN:171115067 @SQ SN:7 LN:159138663 @SQ SN:8 LN:146364022 @SQ SN:9 LN:141213431 @SQ SN:MT LN:16569 @SQ SN:X LN:155270560 @SQ SN:Y LN:59373566 @SQ SN:GL000192.1 LN:547496 @SQ SN:GL000225.1 LN:211173 @SQ SN:GL000194.1 LN:191469 @SQ SN:GL000193.1 LN:189789 @SQ SN:GL000200.1 LN:187035 @SQ SN:GL000222.1 LN:186861 @SQ SN:GL000212.1 LN:186858 @SQ SN:GL000195.1 LN:182896 @SQ SN:GL000223.1 LN:180455 @SQ SN:GL000224.1 LN:179693 @SQ SN:GL000219.1 LN:179198 @SQ SN:GL000205.1 LN:174588 @SQ SN:GL000215.1 LN:172545 @SQ SN:GL000216.1 LN:172294 @SQ SN:GL000217.1 LN:172149 @SQ SN:GL000199.1 LN:169874 @SQ SN:GL000211.1 LN:166566 @SQ SN:GL000213.1 LN:164239 @SQ SN:GL000220.1 LN:161802 @SQ SN:GL000218.1 LN:161147 @SQ SN:GL000209.1 LN:159169 @SQ SN:GL000221.1 LN:155397 @SQ SN:GL000214.1 LN:137718 @SQ SN:GL000228.1 LN:129120 @SQ SN:GL000227.1 LN:128374 @SQ SN:GL000191.1 LN:106433 @SQ SN:GL000208.1 LN:92689 @SQ SN:GL000198.1 LN:90085 @SQ SN:GL000204.1 LN:81310 @SQ SN:GL000233.1 LN:45941 @SQ SN:GL000237.1 LN:45867 @SQ SN:GL000230.1 LN:43691 @SQ SN:GL000242.1 LN:43523 @SQ SN:GL000243.1 LN:43341 @SQ SN:GL000241.1 LN:42152 @SQ SN:GL000236.1 LN:41934 @SQ SN:GL000240.1 LN:41933 @SQ SN:GL000206.1 LN:41001 @SQ SN:GL000232.1 LN:40652 @SQ SN:GL000234.1 LN:40531 @SQ SN:GL000202.1 LN:40103 @SQ SN:GL000238.1 LN:39939 @SQ SN:GL000244.1 LN:39929 @SQ SN:GL000248.1 LN:39786 @SQ SN:GL000196.1 LN:38914 @SQ SN:GL000249.1 LN:38502 @SQ SN:GL000246.1 LN:38154 @SQ SN:GL000203.1 LN:37498 @SQ SN:GL000197.1 LN:37175 @SQ SN:GL000245.1 LN:36651 @SQ SN:GL000247.1 LN:36422 @SQ SN:GL000201.1 LN:36148 @SQ SN:GL000235.1 LN:34474 @SQ SN:GL000239.1 LN:33824 @SQ SN:GL000210.1 LN:27682 @SQ SN:GL000231.1 LN:27386 @SQ SN:GL000229.1 LN:19913 @SQ SN:GL000226.1 LN:15008 @SQ SN:GL000207.1 LN:4262 @PG ID:bowtie2 PN:bowtie2 VN:2.5.1 CL:"/home/xinwei/bin/bowtie2-2.5.1-linux-x86_64/bowtie2-align-s --wrapper basic-0 -t -p 40 -x /home/xinwei/bowtie2_idx_GRCh37/GRCh37 -S /on2/bowtie2_results/IC009T/IC009T.bam -1 /on3/curieT/IC009T/01.fastq.gz,/on3/curieT/IC009T/03.fastq.gz,/on3/curieT/IC009T/05.fastq.gz,/on3/curieT/IC009T/07.fastq.gz,/on3/curieT/IC009T/09.fastq.gz,/on3/curieT/IC009T/11.fastq.gz,/on3/curieT/IC009T/13.fastq.gz,/on3/curieT/IC009T/15.fastq.gz,/on3/curieT/IC009T/17.fastq.gz,/on3/curieT/IC009T/19.fastq.gz,/on3/curieT/IC009T/21.fastq.gz,/on3/curieT/IC009T/23.fastq.gz,/on3/curieT/IC009T/25.fastq.gz -2 /on3/curieT/IC009T/02.fastq.gz,/on3/curieT/IC009T/04.fastq.gz,/on3/curieT/IC009T/06.fastq.gz,/on3/curieT/IC009T/08.fastq.gz,/on3/curieT/IC009T/10.fastq.gz,/on3/curieT/IC009T/12.fastq.gz,/on3/curieT/IC009T/14.fastq.gz,/on3/curieT/IC009T/16.fastq.gz,/on3/curieT/IC009T/18.fastq.gz,/on3/curieT/IC009T/20.fastq.gz,/on3/curieT/IC009T/22.fastq.gz,/on3/curieT/IC009T/24.fastq.gz,/on3/curieT/IC009T/26.fastq.gz" @PG ID:samtools PN:samtools PP:bowtie2 VN:1.16.1 CL:samtools view -@ 40 -bS /on2/bowtie2_results/IC009T/IC009T.sam @PG ID:samtools.1 PN:samtools PP:samtools VN:1.16.1 CL:samtools sort -@ 40 /on2/bowtie2_results/IC009T/IC009T.bam @PG ID:samtools.2 PN:samtools PP:samtools.1 VN:1.16.1 CL:samtools view -H IC009T.sort.bam

GRCh38 samtools view -H IC009T.sort.bam @HD VN:1.5 SO:coordinate @SQ SN:1 LN:248956422 @SQ SN:10 LN:133797422 @SQ SN:11 LN:135086622 @SQ SN:12 LN:133275309 @SQ SN:13 LN:114364328 @SQ SN:14 LN:107043718 @SQ SN:15 LN:101991189 @SQ SN:16 LN:90338345 @SQ SN:17 LN:83257441 @SQ SN:18 LN:80373285 @SQ SN:19 LN:58617616 @SQ SN:2 LN:242193529 @SQ SN:20 LN:64444167 @SQ SN:21 LN:46709983 @SQ SN:22 LN:50818468 @SQ SN:3 LN:198295559 @SQ SN:4 LN:190214555 @SQ SN:5 LN:181538259 @SQ SN:6 LN:170805979 @SQ SN:7 LN:159345973 @SQ SN:8 LN:145138636 @SQ SN:9 LN:138394717 @SQ SN:MT LN:16569 @SQ SN:X LN:156040895 @SQ SN:Y LN:57227415 @SQ SN:KI270728.1 LN:1872759 @SQ SN:KI270727.1 LN:448248 @SQ SN:KI270442.1 LN:392061 @SQ SN:KI270729.1 LN:280839 @SQ SN:GL000225.1 LN:211173 @SQ SN:KI270743.1 LN:210658 @SQ SN:GL000008.2 LN:209709 @SQ SN:GL000009.2 LN:201709 @SQ SN:KI270747.1 LN:198735 @SQ SN:KI270722.1 LN:194050 @SQ SN:GL000194.1 LN:191469 @SQ SN:KI270742.1 LN:186739 @SQ SN:GL000205.2 LN:185591 @SQ SN:GL000195.1 LN:182896 @SQ SN:KI270736.1 LN:181920 @SQ SN:KI270733.1 LN:179772 @SQ SN:GL000224.1 LN:179693 @SQ SN:GL000219.1 LN:179198 @SQ SN:KI270719.1 LN:176845 @SQ SN:GL000216.2 LN:176608 @SQ SN:KI270712.1 LN:176043 @SQ SN:KI270706.1 LN:175055 @SQ SN:KI270725.1 LN:172810 @SQ SN:KI270744.1 LN:168472 @SQ SN:KI270734.1 LN:165050 @SQ SN:GL000213.1 LN:164239 @SQ SN:GL000220.1 LN:161802 @SQ SN:KI270715.1 LN:161471 @SQ SN:GL000218.1 LN:161147 @SQ SN:KI270749.1 LN:158759 @SQ SN:KI270741.1 LN:157432 @SQ SN:GL000221.1 LN:155397 @SQ SN:KI270716.1 LN:153799 @SQ SN:KI270731.1 LN:150754 @SQ SN:KI270751.1 LN:150742 @SQ SN:KI270750.1 LN:148850 @SQ SN:KI270519.1 LN:138126 @SQ SN:GL000214.1 LN:137718 @SQ SN:KI270708.1 LN:127682 @SQ SN:KI270730.1 LN:112551 @SQ SN:KI270438.1 LN:112505 @SQ SN:KI270737.1 LN:103838 @SQ SN:KI270721.1 LN:100316 @SQ SN:KI270738.1 LN:99375 @SQ SN:KI270748.1 LN:93321 @SQ SN:KI270435.1 LN:92983 @SQ SN:GL000208.1 LN:92689 @SQ SN:KI270538.1 LN:91309 @SQ SN:KI270756.1 LN:79590 @SQ SN:KI270739.1 LN:73985 @SQ SN:KI270757.1 LN:71251 @SQ SN:KI270709.1 LN:66860 @SQ SN:KI270746.1 LN:66486 @SQ SN:KI270753.1 LN:62944 @SQ SN:KI270589.1 LN:44474 @SQ SN:KI270726.1 LN:43739 @SQ SN:KI270735.1 LN:42811 @SQ SN:KI270711.1 LN:42210 @SQ SN:KI270745.1 LN:41891 @SQ SN:KI270714.1 LN:41717 @SQ SN:KI270732.1 LN:41543 @SQ SN:KI270713.1 LN:40745 @SQ SN:KI270754.1 LN:40191 @SQ SN:KI270710.1 LN:40176 @SQ SN:KI270717.1 LN:40062 @SQ SN:KI270724.1 LN:39555 @SQ SN:KI270720.1 LN:39050 @SQ SN:KI270723.1 LN:38115 @SQ SN:KI270718.1 LN:38054 @SQ SN:KI270317.1 LN:37690 @SQ SN:KI270740.1 LN:37240 @SQ SN:KI270755.1 LN:36723 @SQ SN:KI270707.1 LN:32032 @SQ SN:KI270579.1 LN:31033 @SQ SN:KI270752.1 LN:27745 @SQ SN:KI270512.1 LN:22689 @SQ SN:KI270322.1 LN:21476 @SQ SN:GL000226.1 LN:15008 @SQ SN:KI270311.1 LN:12399 @SQ SN:KI270366.1 LN:8320 @SQ SN:KI270511.1 LN:8127 @SQ SN:KI270448.1 LN:7992 @SQ SN:KI270521.1 LN:7642 @SQ SN:KI270581.1 LN:7046 @SQ SN:KI270582.1 LN:6504 @SQ SN:KI270515.1 LN:6361 @SQ SN:KI270588.1 LN:6158 @SQ SN:KI270591.1 LN:5796 @SQ SN:KI270522.1 LN:5674 @SQ SN:KI270507.1 LN:5353 @SQ SN:KI270590.1 LN:4685 @SQ SN:KI270584.1 LN:4513 @SQ SN:KI270320.1 LN:4416 @SQ SN:KI270382.1 LN:4215 @SQ SN:KI270468.1 LN:4055 @SQ SN:KI270467.1 LN:3920 @SQ SN:KI270362.1 LN:3530 @SQ SN:KI270517.1 LN:3253 @SQ SN:KI270593.1 LN:3041 @SQ SN:KI270528.1 LN:2983 @SQ SN:KI270587.1 LN:2969 @SQ SN:KI270364.1 LN:2855 @SQ SN:KI270371.1 LN:2805 @SQ SN:KI270333.1 LN:2699 @SQ SN:KI270374.1 LN:2656 @SQ SN:KI270411.1 LN:2646 @SQ SN:KI270414.1 LN:2489 @SQ SN:KI270510.1 LN:2415 @SQ SN:KI270390.1 LN:2387 @SQ SN:KI270375.1 LN:2378 @SQ SN:KI270420.1 LN:2321 @SQ SN:KI270509.1 LN:2318 @SQ SN:KI270315.1 LN:2276 @SQ SN:KI270302.1 LN:2274 @SQ SN:KI270518.1 LN:2186 @SQ SN:KI270530.1 LN:2168 @SQ SN:KI270304.1 LN:2165 @SQ SN:KI270418.1 LN:2145 @SQ SN:KI270424.1 LN:2140 @SQ SN:KI270417.1 LN:2043 @SQ SN:KI270508.1 LN:1951 @SQ SN:KI270303.1 LN:1942 @SQ SN:KI270381.1 LN:1930 @SQ SN:KI270529.1 LN:1899 @SQ SN:KI270425.1 LN:1884 @SQ SN:KI270396.1 LN:1880 @SQ SN:KI270363.1 LN:1803 @SQ SN:KI270386.1 LN:1788 @SQ SN:KI270465.1 LN:1774 @SQ SN:KI270383.1 LN:1750 @SQ SN:KI270384.1 LN:1658 @SQ SN:KI270330.1 LN:1652 @SQ SN:KI270372.1 LN:1650 @SQ SN:KI270548.1 LN:1599 @SQ SN:KI270580.1 LN:1553 @SQ SN:KI270387.1 LN:1537 @SQ SN:KI270391.1 LN:1484 @SQ SN:KI270305.1 LN:1472 @SQ SN:KI270373.1 LN:1451 @SQ SN:KI270422.1 LN:1445 @SQ SN:KI270316.1 LN:1444 @SQ SN:KI270340.1 LN:1428 @SQ SN:KI270338.1 LN:1428 @SQ SN:KI270583.1 LN:1400 @SQ SN:KI270334.1 LN:1368 @SQ SN:KI270429.1 LN:1361 @SQ SN:KI270393.1 LN:1308 @SQ SN:KI270516.1 LN:1300 @SQ SN:KI270389.1 LN:1298 @SQ SN:KI270466.1 LN:1233 @SQ SN:KI270388.1 LN:1216 @SQ SN:KI270544.1 LN:1202 @SQ SN:KI270310.1 LN:1201 @SQ SN:KI270412.1 LN:1179 @SQ SN:KI270395.1 LN:1143 @SQ SN:KI270376.1 LN:1136 @SQ SN:KI270337.1 LN:1121 @SQ SN:KI270335.1 LN:1048 @SQ SN:KI270378.1 LN:1048 @SQ SN:KI270379.1 LN:1045 @SQ SN:KI270329.1 LN:1040 @SQ SN:KI270419.1 LN:1029 @SQ SN:KI270336.1 LN:1026 @SQ SN:KI270312.1 LN:998 @SQ SN:KI270539.1 LN:993 @SQ SN:KI270385.1 LN:990 @SQ SN:KI270423.1 LN:981 @SQ SN:KI270392.1 LN:971 @SQ SN:KI270394.1 LN:970 @PG ID:bowtie2 PN:bowtie2 VN:2.5.1 CL:"/home/xinwei/bin/bowtie2-2.5.1-linux-x86_64/bowtie2-align-s --wrapper basic-0 -t -p 40 -x /home/xinwei/bowtie2_idx_GRCh38/GRCh38 -S /on2/bowtie2_results/IC009T.sam -1 /on3/curieT/IC009T/01.fastq.gz,/on3/curieT/IC009T/03.fastq.gz,/on3/curieT/IC009T/05.fastq.gz,/on3/curieT/IC009T/07.fastq.gz,/on3/curieT/IC009T/09.fastq.gz,/on3/curieT/IC009T/11.fastq.gz,/on3/curieT/IC009T/13.fastq.gz,/on3/curieT/IC009T/15.fastq.gz,/on3/curieT/IC009T/17.fastq.gz,/on3/curieT/IC009T/19.fastq.gz,/on3/curieT/IC009T/21.fastq.gz,/on3/curieT/IC009T/23.fastq.gz,/on3/curieT/IC009T/25.fastq.gz -2 /on3/curieT/IC009T/02.fastq.gz,/on3/curieT/IC009T/04.fastq.gz,/on3/curieT/IC009T/06.fastq.gz,/on3/curieT/IC009T/08.fastq.gz,/on3/curieT/IC009T/10.fastq.gz,/on3/curieT/IC009T/12.fastq.gz,/on3/curieT/IC009T/14.fastq.gz,/on3/curieT/IC009T/16.fastq.gz,/on3/curieT/IC009T/18.fastq.gz,/on3/curieT/IC009T/20.fastq.gz,/on3/curieT/IC009T/22.fastq.gz,/on3/curieT/IC009T/24.fastq.gz,/on3/curieT/IC009T/26.fastq.gz" @PG ID:samtools PN:samtools PP:bowtie2 VN:1.16.1 CL:samtools view -bS -@ 40 IC009T.sam @PG ID:samtools.1 PN:samtools PP:samtools VN:1.16.1 CL:samtools sort -@ 40 IC009T.bam @PG ID:samtools.2 PN:samtools PP:samtools.1 VN:1.16.1 CL:samtools view -H IC009T.sort.bam