Problem with "missing pair end parameters:mean stdev"

[nelsonchanhk]

Hello, I have several BAMs from the same capture and pre-processing was the same for all of them. For some of the BAMs, they are fine with smoove but for some of them the follow error comes up. May I what's the problem? Thanks a lot!

[smoove] 2019/05/04 20:32:56 starting with version 0.2.3 [smoove] 2019/05/04 20:32:56 calculating bam stats for 1 bams [smoove]: ([E]lumpy-filter) 2019/05/04 20:33:01 [lumpy_filter] extracted splits and discordants from 2581973 total aligned reads [smoove]:2019/05/04 20:33:01 finished process: lumpy-filter (set -eu; lumpy_filter -f /home/nelson/OPERATION/RedCellNGS/ref/Homo_sapiens_assembly38.fasta /home/n) in user-time:8.934404s system-time:1.509696s [smoove] 2019/05/04 20:33:02 done calculating bam stats [smoove] 2019/05/04 20:33:18 removed 8807 alignments out of 39644 (22.22%) with low mapq, depth > 1000, or from excluded chroms from 16H28412-3.split.bam in 16 seconds [smoove] 2019/05/04 20:33:18 removed 2587 alignments out of 39644 (6.53%) that were bad interchromosomals or flanked-splitters from 16H28412-3.split.bam [smoove] 2019/05/04 20:33:19 removed 24 singletons of 28250 reads (0.08%) from 16H28412-3.split.bam in 0 seconds [smoove] 2019/05/04 20:33:19 28226 reads (71.20%) of the original 39644 remain from 16H28412-3.split.bam [smoove] 2019/05/04 20:33:28 removed 61609 alignments out of 2554480 (2.41%) with low mapq, depth > 1000, or from excluded chroms from 16H28412-3.disc.bam in 25 seconds [smoove] 2019/05/04 20:33:28 removed 1 alignments out of 2554480 (0.00%) that were bad interchromosomals or flanked-splitters from 16H28412-3.disc.bam [smoove] 2019/05/04 20:33:41 removed 68 singletons and isolated interchromosomals of 2492870 reads (0.00%) from 16H28412-3.disc.bam in 13 seconds [smoove] 2019/05/04 20:33:41 2492802 reads (97.59%) of the original 2554480 remain from 16H28412-3.disc.bam [smoove] 2019/05/04 20:33:41 starting lumpy [smoove] 2019/05/04 20:33:41 wrote lumpy command to /home/nelson/OPERATION/RedCellNGS/Processed_bam_vcf_1/smoove/16H28412-3-lumpy-cmd.sh [smoove] 2019/05/04 20:33:41 writing sorted, indexed file to /home/nelson/OPERATION/RedCellNGS/Processed_bam_vcf_1/smoove/16H28412-3-smoove.genotyped.vcf.gz [smoove] 2019/05/04 20:33:41 excluding variants with all unknown or homozygous reference genotypes [smoove] 2019/05/04 20:33:41 missing pair end parameters:mean stdev

[smoove] 2019/05/04 20:33:41 Program: ********** (v 0.2.13) Author: Ryan Layer ([email protected]) Summary: Find structural variations in various signals.

Usage: ********** [OPTIONS]

Options: -g Genome file (defines chromosome order) -e Show evidence for each call -w File read windows size (default 1000000) -mw minimum weight for a call -msw minimum per-sample weight for a call -tt trim threshold -x exclude file bed file -t temp file prefix, must be to a writeable directory -P output probability curve for each variant -b output BEDPE instead of VCF -sr bam_file:, id:, back_distance:, min_mapping_threshold:, [smoove] 2019/05/04 20:33:41 weight:, min_clip:, read_group:

-pe	bam_file:<file name>,
	id:<sample name>,
	histo_file:<file name>,
	mean:<value>,
	stdev:<value>,

[smoove] 2019/05/04 20:33:41 read_length:, min_non_overlap:, discordant_z:, back_distance:, min_mapping_threshold:, [smoove] 2019/05/04 20:33:41 weight:, read_group:

[smoove] 2019/05/04 20:33:41 -bedpe bedpe_file:, id:, weight:

[smoove] 2019/05/04 20:33:41 > gsort version 0.0.6 [smoove] 2019/05/04 20:33:48 2019/05/04 20:33:48 EOF [smoove] 2019/05/04 20:33:48 Failed to open -: unknown file type [smoove] 2019/05/04 20:33:48 Failed to open -: unknown file type panic: exit status 255

goroutine 1 [running]: github.com/brentp/smoove/svtyper.check(...) /home/brentp/go/src/github.com/brentp/smoove/svtyper/svtyper.go:33 github.com/brentp/smoove/svtyper.Svtyper(0xa88960, 0xc0001bc140, 0x7fff3beabed6, 0x43, 0xc000190670, 0x1, 0x1, 0x7fff3beabe91, 0x3c, 0x7fff3beabe7d, ...) /home/brentp/go/src/github.com/brentp/smoove/svtyper/svtyper.go:226 +0x18c5 github.com/brentp/smoove/lumpy.Main() /home/brentp/go/src/github.com/brentp/smoove/lumpy/lumpy.go:346 +0x468 main.main() /home/brentp/go/src/github.com/brentp/smoove/cmd/smoove/smoove.go:124 +0x1d7

[brentp]

what is the content of /home/nelson/OPERATION/RedCellNGS/Processed_bam_vcf_1/smoove/16H28412-3-lumpy-cmd.sh?

[nelsonchanhk]

Hello brentp, thanks for your quick reply. It's as follows. Thanks!

set -euo pipefail set -euo pipefail; lumpy -msw 4 -mw 4 -t $(mktemp) -tt 0 -P -pe id:16H28412-3,bam_file:/home/nelson/OPERATION/RedCellNGS/Processed_bam_vcf_1/smoove/16H28412-3.disc.bam,histo_file:/home/nelson/OPERATION/RedCellNGS/Processed_bam_vcf_1/smoove/16H28412-3.histo,mean:0.00,stdev:0.00,read_length:151,min_non_overlap:151,discordant_z:4,back_distance:30,weight:1,min_mapping_threshold:20 -sr id:16H28412-3,bam_file:/home/nelson/OPERATION/RedCellNGS/Processed_bam_vcf_1/smoove/16H28412-3.split.bam,back_distance:10,weight:1,min_mapping_threshold:20

[brentp]

something is wrong with that bam file, note how it has: mean:0.00,stdev:0.00

I would do a samtools flagstat on the bam and do a samtools view and look and see if reads are properly paired and what the insert sizes are.

[nelsonchanhk]

Hello Brent, it's the problem of my bam, thanks for your expert assistance! Have a nice weekend!