Cannot extract any read from IGSR High Coverage sample

[NTNguyen13]

Hi, I tried Kourami for HLA typing recently, the tool itself and the preprocessing script run fine on my sample (aligned on Hg38 + decoy + HLA + no ALT). However, when I tried it on IGSR high coverage sample (http://ftp.1000genomes.ebi.ac.uk/vol1/ftp/data_collections/1000G_2504_high_coverage/), the preprocessing script return zero reads. As far as I known, the sample is aligned on Hg38 + decoy + HLA + ALT, but I tried all four scripts, but they all return zero read (I checked the file manually)

Have you ever get into this problem? Please give me some insight on this.

Here is the running log of the preprocessing script.

sample_name=HG02082.final
file=/home/nguyen/IGSR_HC_Data/CRAM/HG02082.final.cram

(base) nguyen@nguyen-ws:~$     $KOUDIR/scripts/alignAndExtract_hs38DH.sh $out_folder/${sample_name}/kourami/$sample_name $file
>>>>>>>>>>>>>>>> extracting reads mapping to HLA loci and ALT contigs (38DH)
[main_samview] random alignment retrieval only works for indexed BAM or CRAM files.
>>>>>>>>>>>>>> indexing extracted bam (38DH)
>>>>>>>>>>>>>> bamUtil fastq extraction (38DH)
>>>>>>>>>>>>>> Mapping back to GRCh38_NoALT_wHLA (38DH_NoAlt)
[M::bwa_idx_load_from_disk] read 0 ALT contigs
[main] Version: 0.7.17-r1188
[main] CMD: /home/nguyen/anaconda3/bin/bwa mem -t 8 /home/nguyen/Exec/kourami-0.9.6/scripts/../resources/hs38NoAltDH.fa /home/nguyen/IGSR_HC_Data/Result/HG02082.final/kourami/HG02082.final_tmp.extract_1.fq.gz /home/nguyen/IGSR_HC_Data/Result/HG02082.final/kourami/HG02082.final_tmp.extract_2.fq.gz
[main] Real time: 1.905 sec; CPU: 1.908 sec
>>>>>>>>>>>>>> indexing extracted bam (38DH_NoAlt)
>>>>>>>>>>>>>> bamUtil fastq extraction (38DH_NoAlt)
>>>>>>>>>>>>>> bwa mem to hla panel for Kourami
[M::bwa_idx_load_from_disk] read 0 ALT contigs
[main] Version: 0.7.17-r1188
[main] CMD: /home/nguyen/anaconda3/bin/bwa mem -t 8 /home/nguyen/Exec/kourami-0.9.6/scripts/../db/All_FINAL_with_Decoy.fa.gz /home/nguyen/IGSR_HC_Data/Result/HG02082.final/kourami/HG02082.final_extract_1.fq.gz /home/nguyen/IGSR_HC_Data/Result/HG02082.final/kourami/HG02082.final_extract_2.fq.gz
[main] Real time: 0.056 sec; CPU: 0.056 sec

(base) nguyen@nguyen-ws:~$     $KOUDIR/scripts/alignAndExtract_hs38DH_NoAlt.sh $out_folder/${sample_name}/kourami/$sample_name $file
>>>>>>>>>>>>>>>> extracting reads mapping to HLA loci and ALT contigs (38DH_NoAlt)
[main_samview] random alignment retrieval only works for indexed BAM or CRAM files.
>>>>>>>>>>>>>> indexing extracted bam (38DH_NoAlt)
>>>>>>>>>>>>>> bamUtil fastq extraction (38DH_NoAlt)
>>>>>>>>>>>>>> bwa mem to hla panel for Kourami
[M::bwa_idx_load_from_disk] read 0 ALT contigs
[main] Version: 0.7.17-r1188
[main] CMD: /home/nguyen/anaconda3/bin/bwa mem -t 8 /home/nguyen/Exec/kourami-0.9.6/scripts/../db/All_FINAL_with_Decoy.fa.gz /home/nguyen/IGSR_HC_Data/Result/HG02082.final/kourami/HG02082.final_extract_1.fq.gz /home/nguyen/IGSR_HC_Data/Result/HG02082.final/kourami/HG02082.final_extract_2.fq.gz
[main] Real time: 0.056 sec; CPU: 0.057 sec

(base) nguyen@nguyen-ws:~$     $KOUDIR/scripts/alignAndExtract_hs38Alt.sh $out_folder/${sample_name}/kourami/$sample_name $file
>>>>>>>>>>>>>>>> extracting reads mapping to HLA loci and ALT contigs (38Alt)
samtools view: Could not read file "/home/nguyen/Exec/kourami-0.9.6/scripts/../resources/hs38dh.initial.bed": No such file or directory
[main_samview] random alignment retrieval only works for indexed BAM or CRAM files.
>>>>>>>>>>>>>> indexing extracted bam (38Alt)
>>>>>>>>>>>>>> bamUtil fastq extraction (38Alt)
rm /home/nguyen/IGSR_HC_Data/Result/HG02082.final/kourami/HG02082.final.tmp.extract.bam /home/nguyen/IGSR_HC_Data/Result/HG02082.final/kourami/HG02082.final.tmp.extract.bam.bai
>>>>>>>>>>>>>> Mapping back to GRCh38_NoALT_wHLA (38DH_NoAlt)
[M::bwa_idx_load_from_disk] read 0 ALT contigs
[main] Version: 0.7.17-r1188
[main] CMD: /home/nguyen/anaconda3/bin/bwa mem -t 8 /home/nguyen/Exec/kourami-0.9.6/scripts/../resources/hs38NoAltDH.fa /home/nguyen/IGSR_HC_Data/Result/HG02082.final/kourami/HG02082.final_tmp.extract_1.fq.gz /home/nguyen/IGSR_HC_Data/Result/HG02082.final/kourami/HG02082.final_tmp.extract_2.fq.gz
[main] Real time: 34.010 sec; CPU: 8.334 sec
>>>>>>>>>>>>>> indexing extracted bam (38DH_NoAlt)
>>>>>>>>>>>>>> bamUtil fastq extraction (38DH_NoAlt)
>>>>>>>>>>>>>> bwa mem to hla panel for Kourami
[M::bwa_idx_load_from_disk] read 0 ALT contigs
[main] Version: 0.7.17-r1188
[main] CMD: /home/nguyen/anaconda3/bin/bwa mem -t 8 /home/nguyen/Exec/kourami-0.9.6/scripts/../db/All_FINAL_with_Decoy.fa.gz /home/nguyen/IGSR_HC_Data/Result/HG02082.final/kourami/HG02082.final_extract_1.fq.gz /home/nguyen/IGSR_HC_Data/Result/HG02082.final/kourami/HG02082.final_extract_2.fq.gz
[main] Real time: 0.068 sec; CPU: 0.069 sec

(base) nguyen@nguyen-ws:~$ $KOUDIR/scripts/alignAndExtract_hs38.sh $out_folder/${sample_name}/kourami/$sample_name $file
>>>>>>>>>>>>>>>> extracting reads mapping to HLA loci and ALT contigs (38)
[main_samview] random alignment retrieval only works for indexed BAM or CRAM files.
>>>>>>>>>>>>>> indexing extracted bam (38)
>>>>>>>>>>>>>> bamUtil fastq extraction (38)
rm /home/nguyen/IGSR_HC_Data/Result/HG02082.final/kourami/HG02082.final.tmp.extract.bam /home/nguyen/IGSR_HC_Data/Result/HG02082.final/kourami/HG02082.final.tmp.extract.bam.bai
>>>>>>>>>>>>>> Mapping back to GRCh38_NoALT_wHLA (38DH_NoAlt)
[M::bwa_idx_load_from_disk] read 0 ALT contigs
[main] Version: 0.7.17-r1188
[main] CMD: /home/nguyen/anaconda3/bin/bwa mem -t 8 /home/nguyen/Exec/kourami-0.9.6/scripts/../resources/hs38NoAltDH.fa /home/nguyen/IGSR_HC_Data/Result/HG02082.final/kourami/HG02082.final_tmp.extract_1.fq.gz /home/nguyen/IGSR_HC_Data/Result/HG02082.final/kourami/HG02082.final_tmp.extract_2.fq.gz
[main] Real time: 1.898 sec; CPU: 1.899 sec
>>>>>>>>>>>>>> indexing extracted bam (38DH_NoAlt)
>>>>>>>>>>>>>> bamUtil fastq extraction (38DH_NoAlt)
>>>>>>>>>>>>>> bwa mem to hla panel for Kourami
[M::bwa_idx_load_from_disk] read 0 ALT contigs
[main] Version: 0.7.17-r1188
[main] CMD: /home/nguyen/anaconda3/bin/bwa mem -t 8 /home/nguyen/Exec/kourami-0.9.6/scripts/../db/All_FINAL_with_Decoy.fa.gz /home/nguyen/IGSR_HC_Data/Result/HG02082.final/kourami/HG02082.final_extract_1.fq.gz /home/nguyen/IGSR_HC_Data/Result/HG02082.final/kourami/HG02082.final_extract_2.fq.gz
[main] Real time: 0.059 sec; CPU: 0.061 sec