BR: HMMRATAC fails due to SAM validation error?

[hisplan]

Describe the bug

HMMRATAC fails with this error message:

Exception in thread "main" net.sf.samtools.SAMFormatException: SAM validation error: ERROR: Record 292169, Read name A00519:269:H7FM2DRXX:1:1218:5303:31219, Zero-length read without FZ, CS or CQ tag
	at net.sf.samtools.SAMUtils.processValidationErrors(SAMUtils.java:448)
	at net.sf.samtools.BAMFileReader$BAMFileIterator.advance(BAMFileReader.java:541)
	at net.sf.samtools.BAMFileReader$BAMFileIterator.next(BAMFileReader.java:522)
	at net.sf.samtools.BAMFileReader$BAMFileIterator.next(BAMFileReader.java:481)
	at net.sf.samtools.BAMFileReader$BAMQueryFilteringIterator.advance(BAMFileReader.java:750)
	at net.sf.samtools.BAMFileReader$BAMQueryFilteringIterator.next(BAMFileReader.java:720)
	at net.sf.samtools.BAMFileReader$BAMQueryFilteringIterator.next(BAMFileReader.java:673)
	at net.sf.samtools.SAMFileReader$AssertableIterator.next(SAMFileReader.java:672)
	at net.sf.samtools.SAMFileReader$AssertableIterator.next(SAMFileReader.java:650)
	at WigMath.pileup.makeBlock(pileup.java:228)
	at WigMath.pileup.build(pileup.java:205)
	at WigMath.pileup.<init>(pileup.java:70)
	at HMMR_ATAC.Main_HMMR_Driver.main(Main_HMMR_Driver.java:270)

Here is the actual reads that HMMRATAC complained about:

$ samtools view possorted_bam.bam | grep -F "A00519:269:H7FM2DRXX:1:1218:5303:31219"
A00519:269:H7FM2DRXX:1:1218:5303:31219	69	chr6	148759869	0	*	=	148759869	0	*	*	MC:Z:12S37M	AS:i:0	XS:i:0	CR:Z:GGTGAAGCACTGTCGG	CY:Z:FF:F,FFFFFFFFFFF	CB:Z:GGTGAAGCACTGTCGG-1	BC:Z:CTTTCGAC	QT:Z::FFFFFFF	TR:Z:TGTCTCTTATACACATCTCCGAGCCCACGAGACCTTTCGACATCTCGTAT	TQ:Z:FFFFFFFFF:FFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFF,:FFFF	GP:i:-1	MP:i:1211301816	MQ:i:22	RG:Z:atac_pbmc_500_v1:MissingLibrary:1:H7FM2DRXX:1
A00519:269:H7FM2DRXX:1:1218:5303:31219	137	chr6	148759869	22	12S37M	=	148759869	0	GGGGGGGGTTTTAAAAAAAAAAAAAACCAAAACAAACAAAAAAAAAAAC	FF,FF:,:,,F,,,,,:F:::F,,,,,F:,,:,,,:,F,:,:,,,F::,	NM:i:0	MD:Z:37	AS:i:37	XS:i:32	XA:Z:chr13,+32718338,17S32M,0;chr3,-57922436,5S32M12S,0;chrX,-147034285,5S32M12S,0;chr3,-184460874,2S35M12S,1;chr20,-52312303,9S30M10S,0;	CR:Z:GGTGAAGCACTGTCGG	CY:Z:FF:F,FFFFFFFFFFF	CB:Z:GGTGAAGCACTGTCGG-1	BC:Z:CTTTCGAC	QT:Z::FFFFFFF	GP:i:1211301816	MP:i:-1	MQ:i:0	RG:Z:atac_pbmc_500_v1:MissingLibrary:1:H7FM2DRXX:1

To Reproduce

Only using these absolutely mandatory parameters: --bam, --index, --genome, and --output. Nothing else.

The dataset is 10x's public datasets: 500 PBMC from a healthy donor.

System (please complete the following information):

• Open JDK 8
• HMMRATAC v1.2.9

Thanks for your help.

[EvanTarbell]

Thanks for your comment. It seems that this is one of two places where i use samtools to read a bam file. Although in the other place i use a try/catch block to avoid these format errors, i forgot to add such a block to this particular piece of code. i will add that and release a new version that should solve this issue for you. I should be able to do this soon

[hisplan]

Thanks for your prompt response! Looking forward to trying this out.

[EvanTarbell]

Version 1.2.10 should fix this issue. Let me know if it works for you.

[hisplan]

Thanks! Will try out and let you know.

[hisplan]

Thanks. It worked!

[zyaiqy]

Hello, I also have this problem in version 1.2.10, the specific code is as follows:
Exception in thread "main" net.sf.samtools.SAMFormatException: SAM validation error: ERROR: Record 1, Read name A01758:140:HLVFVDSX3:3:2551:23439:35008, Zero-length read without FZ, CS or CQ tag
at net.sf.samtools.SAMUtils.processValidationErrors(SAMUtils.java:448)
at net.sf.samtools.BAMFileReader$BAMFileIterator.advance(BAMFileReader.java:541)
at net.sf.samtools.BAMFileReader$BAMFileIndexIterator.(BAMFileReader.java:648)
at net.sf.samtools.BAMFileReader.createIndexIterator(BAMFileReader.java:598)
at net.sf.samtools.BAMFileReader.query(BAMFileReader.java:352)
at net.sf.samtools.SAMFileReader.query(SAMFileReader.java:363)
at HMMR_ATAC.pullLargeLengths.read(pullLargeLengths.java:112)
at HMMR_ATAC.pullLargeLengths.(pullLargeLengths.java:61)
at HMMR_ATAC.Main_HMMR_Driver.main(Main_HMMR_Driver.java:219)
The following is a specific read of the report:
(samtools) ubuntu@VM-0-12-ubuntu:~/data$ samtools view LJL-6M-1-fin.bam | grep -F "A01758:140:HLVFVDSX3:3:2551:23439:35008"
A01758:140:HLVFVDSX3:3:2551:23439:35008 99 1 583 42 146M = 663 229 * *
A01758:140:HLVFVDSX3:3:2551:23439:35008 147 1 663 42 145M = 583 -229 * *

Thanks for your help.

[wzcc-hub]

Hello, I also have this problem in version 1.2.10, the specific code is as follows: Exception in thread "main" net.sf.samtools.SAMFormatException: SAM validation error: ERROR: Record 1, Read name A01758:140:HLVFVDSX3:3:2551:23439:35008, Zero-length read without FZ, CS or CQ tag at net.sf.samtools.SAMUtils.processValidationErrors(SAMUtils.java:448) at net.sf.samtools.BAMFileReader$BAMFileIterator.advance(BAMFileReader.java:541) at net.sf.samtools.BAMFileReader$BAMFileIndexIterator.(BAMFileReader.java:648) at net.sf.samtools.BAMFileReader.createIndexIterator(BAMFileReader.java:598) at net.sf.samtools.BAMFileReader.query(BAMFileReader.java:352) at net.sf.samtools.SAMFileReader.query(SAMFileReader.java:363) at HMMR_ATAC.pullLargeLengths.read(pullLargeLengths.java:112) at HMMR_ATAC.pullLargeLengths.(pullLargeLengths.java:61) at HMMR_ATAC.Main_HMMR_Driver.main(Main_HMMR_Driver.java:219) The following is a specific read of the report: (samtools) ubuntu@VM-0-12-ubuntu:~/data$ samtools view LJL-6M-1-fin.bam | grep -F "A01758:140:HLVFVDSX3:3:2551:23439:35008" A01758:140:HLVFVDSX3:3:2551:23439:35008 99 1 583 42 146M = 663 229 * * A01758:140:HLVFVDSX3:3:2551:23439:35008 147 1 663 42 145M = 583 -229 * *

Thanks for your help.

have encountered the same problem and I would like to ask if you have solved it?

[simontimmins]

Just to add, I have also come across this same error

[chklee30]

Were you able to figure this out @zyaiqy? I have the exact same error