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I am analyzing an experiment in which paired end sequencing was used to sequence exomes from a number of samples. All of the mapping and bam files were created using lifescope. We ran barcoded samples with multiple samples/lane and multiple lanes/sample. The bam files produced from each lane were merged to create one bam file for each sample. When I run the Picard ValidateSamFile command, I get the following errors:
Mate alignment does not match alignment start of mate
Mate negative strand flag does not match read negative strand flag of mate
Both mates are marked as second of pair
This indicates that the alignments across mate pairs do not coincide. I am concerned about the validity of these alignments. Should they be filtered out prior to downstream processing? Picard has a tool FixMateInformation but I can't find any information on what this tool actually does. I assume it either deletes the offending reads or forces the information across mate pairs to coincide.
Does anyone have more experience with these errors? Can you point me in the right direction?
Thanks,
Mike
Mate alignment does not match alignment start of mate
Mate negative strand flag does not match read negative strand flag of mate
Both mates are marked as second of pair
This indicates that the alignments across mate pairs do not coincide. I am concerned about the validity of these alignments. Should they be filtered out prior to downstream processing? Picard has a tool FixMateInformation but I can't find any information on what this tool actually does. I assume it either deletes the offending reads or forces the information across mate pairs to coincide.
Does anyone have more experience with these errors? Can you point me in the right direction?
Thanks,
Mike
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