HaplotypeCaller ERROR with ERC invalid

Hi everyone,
I'm a really novice on GATK. I want to call SNPs from 12 samples and I got some troubles.
Firstly, I combined 12 samples' bam files using the command "HaplotypeCaller". Each bam file stands for one sample and is generated by two PE sequencing files (R1 & R2). However, I got the ERROR:

A USER ERROR has occurred: Argument --emitRefConfidence has a bad value: Can only be used in single sample mode currently. Use the sample_name argument to run on a single sample out of a multi-sample BAM file.

Refer to the problem before,it seems that the BAM file has multiple lanes message. I checked the SAM file's head lines and tail lines. The head lines are as following:
HWI-7001446:625:C9078ANXX:6:1101:1209:2074 ……
HWI-7001446:625:C9078ANXX:6:1101:1209:2074 ……
HWI-7001446:625:C9078ANXX:6:1101:1232:2088 ……
HWI-7001446:625:C9078ANXX:6:1101:1232:2088 ……
And the tail lines are as following:
ST-E00159:217:HYL2KCCXX:6:2224:23979:72667 ……
ST-E00159:217:HYL2KCCXX:6:2224:23979:72667 ……
ST-E00159:217:HYL2KCCXX:6:2224:24000:72667 ……
ST-E00159:217:HYL2KCCXX:6:2224:24000:72667 ……

Next, I format the BAM file by the command "AddOrReplaceReadGroups". Then I run the HaplotypeCaller command on the formated BAM file. The .g.vcf file has been generated and I got another ERROR:

A USER ERROR has occurred: Traversal by intervals was requested but some input files are not indexed.
Please index all input files:
samtools index xxx.group.bam

Is the generated .g.vcf file valid? Why did I get the two ERROR messages?

Thank u very much.

Best wishes,

Tony

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Best Answer

  • Accepted Answer

    Besides, the formated BAM file is similar to the initial BAM file. I have checked the formated BAM file's SAM file. In fact, it seems that it has been added several lines:
    The 1st line: @HD VN:1.5 SO:unsorted
    The last line before "alignment section": @RG ID:4 LB:lib1 PL:illumina SM:20 PU:unit1
    And other lines look the same……
    The head lines and tail lines are still different as he initial SAM file.

Answers

  • Tony_LeeTony_Lee Member
    Accepted Answer

    Besides, the formated BAM file is similar to the initial BAM file. I have checked the formated BAM file's SAM file. In fact, it seems that it has been added several lines:
    The 1st line: @HD VN:1.5 SO:unsorted
    The last line before "alignment section": @RG ID:4 LB:lib1 PL:illumina SM:20 PU:unit1
    And other lines look the same……
    The head lines and tail lines are still different as he initial SAM file.

  • SheilaSheila Broad InstituteMember, Broadie, Moderator admin

    @Tony_Lee
    Hi Tony,

    The error message suggests your BAM files are not indexed. You need to use samtools index to index them.

    -Sheila

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