toddajohnson
commented
1 year ago If you run samtools view -H COLO829R.bam > check.sam against one of those test bam files, you will see in the output header that the reference genome used for mapping is "/opt/resources/reference_genome/37/Homo_sapiens.GRCh37.GATK.illumina.fasta"; so, that test data was mapped to build 37 instead of 38, but you are telling amber to use 38 and using build 38 GermlineHetPon. Although I presume that your own data will be mapped will be mapped to the more recent GRCh38, to practice with this test data, you will need to download the build 37 pipeline reference data. The reference used by the test data is mentioned on "https://github.com/hartwigmedical/hmftools/blob/master/pipeline/README_WGS.md" as being GRCh37 under the "Test Data" section. I guess that HMF has lots of people use their resources bandwidth just to download "old" reference data to test the pipeline... :-)
ZWael
p-priestley
Hello I am trying to use AMBER ans I'am not familiar to the java language, I think i have truble with parallelisation and workers here are my steps
1) I created a conda env and installed maven 2) download the amber-3.9.jar (https://github.com/hartwigmedical/hmftools/releases/download/amber-v3.9/amber-3.9.jar) in my local machine 3) download the test files COLO829R.bam and COLO829T.bam from the repo github.com/hartwigmedical/hmftools/tree/master/pipeline/test_data/ 4) download the GermlineHetPon.38.vcf.gz from the nextcloud folder FHMFTools-Resources>Amber>38 j’exécute la commande :
java -Xmx32G -cp AMBER/amber-3.9.jar com.hartwig.hmftools.amber.AmberApplication -reference COLO829R -reference_bam test/COLO829R.bam -tumor COLO829T -tumor_bam test/COLO829T.bam -output_dir out -loci Resources/GermlineHetPon.38.vcf.gz -ref_genome_version 38the output message was13:33:05.454 INFO [main] - AMBER version: 3.9 13:33:05.457 INFO [main] - Loading vcf file Resources/GermlineHetPon.38.vcf.gz 13:33:10.131 INFO [main] - loaded 1337724 baf loci 13:33:10.193 INFO [main] - Processing 1337724 potential sites in reference bam test/COLO829R.bam 13:33:12.436 INFO [main] - 1337724 loci, 148424 genome regions, min gap = 4000 13:33:12.476 INFO [main] - 1 bam reader threads started 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)
13:33:12.483 INFO [main] - 1 bam reader threads finished
13:33:12.984 INFO [main] - Median normal depth is 0 reads: filtering reads outside of 0 and 0
13:33:13.336 INFO [main] - 0 heterozygous, 0 homozygous in reference bams
13:33:13.485 INFO [main] - Median normal depth is 0 reads: filtering reads outside of 0 and 0
13:33:13.527 INFO [main] - Processing 0 germline heterozygous loci in tumor bam test/COLO829T.bam
13:33:13.528 INFO [main] - Processing 0 germline homozygous loci in tumor bam test/COLO829T.bam for contamination
13:33:13.531 INFO [main] - 0 loci, 0 genome regions, min gap = 2000
13:33:13.537 INFO [main] - 1 bam reader threads started
13:33:13.538 INFO [main] - 1 bam reader threads finished
13:33:13.546 INFO [main] - Median tumor depth at potential contamination sites is 0 reads
13:33:13.554 INFO [main] - No evidence of contamination.
13:33:13.581 INFO [main] - Writing 0 contamination records to out/COLO829T.amber.contamination.vcf.gz
13:33:13.690 INFO [main] - Writing 1101 germline snp records to out/COLO829R.amber.snp.vcf.gz
13:33:14.010 INFO [main] - Applying pcf segmentation
13:33:14.020 INFO [main] - Executing R script via command: Rscript /tmp/script2398773644644405398.R out/COLO829T.amber.baf.tsv.gz out/COLO829T.amber.baf.pcf