Downloading: s3://natera-platform-sandbox/pipeline-resources/AIH/rna/GRCh38/starfusion/ctat_genome_lib_build_dir/ref_genome.fa
  Downloading: s3://natera-platform-sandbox/pipeline-resources/Homo_sapiens_assembly38.dbsnp138.vcf.gz
  Downloading: s3://natera-rnd-pltf-dev-nextflow-scratch-01/work/a3/8fc3fc626725ba203f9a0b7ef38667/.command.sh
  Downloading: s3://natera-platform-sandbox/pipeline-resources/AIH/rna/GRCh38/starfusion/ctat_genome_lib_build_dir/ref_genome.dict
  Downloading: s3://natera-rnd-pltf-dev-nextflow-scratch-01/work/6a/b32b913c361db2aaacc6a72613def3/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bam
  Downloading: s3://natera-platform-sandbox/pipeline-resources/AIH/rna/GRCh38/gatk4/dbsnp_hg38_contam_sites.bed
  Downloading: s3://natera-rnd-pltf-dev-nextflow-scratch-01/work/a3/8fc3fc626725ba203f9a0b7ef38667/.command.run
  Downloading: s3://natera-rnd-pltf-dev-nextflow-scratch-01/work/6a/b32b913c361db2aaacc6a72613def3/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
  Downloading: s3://natera-platform-sandbox/pipeline-resources/AIH/rna/GRCh38/starfusion/ctat_genome_lib_build_dir/ref_genome.fa.fai
  Downloading: s3://natera-platform-sandbox/pipeline-resources/beds/twist/Twist_rna_exome_AND_fusion_hg38.bed
  Downloading: s3://natera-platform-sandbox/pipeline-resources/Homo_sapiens_assembly38.dbsnp138.vcf.gz.tbi
==> STAGING COMPLETE (11 inputs)

+ mkdir aih-tih-sc-82e8c1-R1_B23WHYVLT4_1
+ all_qc -bam aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bam -beds Twist_rna_exome_AND_fusion_hg38.bed -dbsnp_vcf Homo_sapiens_assembly38.dbsnp138.vcf.gz -contamination_bed dbsnp_hg38_contam_sites.bed -output_folder aih-tih-sc-82e8c1-R1_B23WHYVLT4_1 -reference ref_genome.fa -region chr20
mkdir -p aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/


cat Twist_rna_exome_AND_fusion_hg38.bed | cut -f 1-3 | bedtools sort -g ref_genome.fa.fai | bedtools merge > aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/full_bed.bed


seqtool depth -bam aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bam -bed aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/full_bed.bed -threads 4 -bam_threads 2 | bgzip -c > aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.depth.gz
saturation_plot       -bams aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bam       -region chr20       -output aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/saturation_plot.png       -threads 1 
seqtool saturation_metric       -at 1000000       -region chr20       -bam_threads 1       aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bam > aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/saturation_qc.tsv 
fragment_size       -bams aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bam       -region chr20       -output_prefix aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/fragsize       -threads 1 
samtools mpileup   -B --min-MQ 60 --min-BQ 30    -f ref_genome.fa   -l dbsnp_hg38_contam_sites.bed   aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bam > aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.mpileup && contamination -input aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.mpileup -dbsnp_vcf Homo_sapiens_assembly38.dbsnp138.vcf.gz > aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/contamination.tsv 
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[mpileup] 1 samples in 1 input files
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
/opt/conda/lib/python3.9/site-packages/pyranges/__init__.py:8: UserWarning: pkg_resources is deprecated as an API. See https://setuptools.pypa.io/en/latest/pkg_resources.html. The pkg_resources package is slated for removal as early as 2025-11-30. Refrain from using this package or pin to Setuptools<81.
  import pkg_resources
/opt/conda/lib/python3.9/site-packages/pyranges/__init__.py:8: UserWarning: pkg_resources is deprecated as an API. See https://setuptools.pypa.io/en/latest/pkg_resources.html. The pkg_resources package is slated for removal as early as 2025-11-30. Refrain from using this package or pin to Setuptools<81.
  import pkg_resources
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai

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[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai




[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai


[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai


tabix -s 1 -b 2 -e 2 aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.depth.gz


seqtool depth_hist     -nbins 500     aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.depth.gz     > aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.depth_hist.tsv


cat aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.depth_hist.tsv | plot         -x 0         -xlabel Depth -ylabel "Number of Sites"         -xlim 0 2000         -title aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md         -output aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.depth.png


[W::hts_idx_load3] The index file is older than the data file: Homo_sapiens_assembly38.dbsnp138.vcf.gz.tbi


seqtool read_qc    -bed Twist_rna_exome_AND_fusion_hg38.bed     -n_threads 1     -bam_threads 2     aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bam > aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/read_qc.Twist_rna_exome_AND_fusion_hg38.bed.tsv
seqtool reads_per_target     -bed Twist_rna_exome_AND_fusion_hg38.bed     -threads 1     -bam_threads 2     aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bam > aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/reads_per_target.Twist_rna_exome_AND_fusion_hg38.bed.tsv 
seqtool depth_stats  -bed Twist_rna_exome_AND_fusion_hg38.bed  aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.depth.gz> aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/depth_stats.Twist_rna_exome_AND_fusion_hg38.bed.tsv 
seqtool gc_bias  -reference ref_genome.fa  -bed Twist_rna_exome_AND_fusion_hg38.bed  -bin_size 0.05  -threads 1  -bam_threads 2  aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bam  > aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/gc_bias_qc.Twist_rna_exome_AND_fusion_hg38.bed.tsv 
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai
[W::hts_idx_load3] The index file is older than the data file: aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md.bai




cat aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/gc_bias_qc.Twist_rna_exome_AND_fusion_hg38.bed.tsv  | plot -x 0  -header infer  -ylim 0 2  -title "aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md" -suptitle "Twist_rna_exome_AND_fusion_hg38.bed" -output "aih-tih-sc-82e8c1-R1_B23WHYVLT4_1/aih-tih-sc-82e8c1-R1_B23WHYVLT4_1.md/gc_bias.Twist_rna_exome_AND_fusion_hg38.bed.png"  -ylabel "Fold enrichment / depletion"  -xlabel "GC Quantile"