SplitNCigarReads results in all reads failing MalformedReadFilter?
Hi, I'm following the GATK best Practices workflow for SNP and indel calling on my RNAseq data, I'm currently at the "Split'N'Trim and reassign mapping qualities" step but come across some problems, below is my command:
java -jar GenomeAnalysisTK.jar -T SplitNCigarReads -R ref.fa -I dedupped.bam -o split.bam -rf \ ReassignOneMappingQuality -RMQF 255 -RMQT 60 -U ALLOW_N_CIGAR_READS \ --filter_mismatching_base_and_quals
Here's what I got:
INFO 00:13:19,415 ProgressMeter - Total runtime 66.03 secs, 1.10 min, 0.02 hours
INFO 00:13:19,418 MicroScheduler - 37770935 reads were filtered out during the traversal out of approximately 37770935 total reads (100.00%)
INFO 00:13:19,418 MicroScheduler - -> 0 reads (0.00% of total) failing BadCigarFilter
INFO 00:13:19,418 MicroScheduler - -> 37770935 reads (100.00% of total) failing MalformedReadFilter
INFO 00:13:19,419 MicroScheduler - -> 0 reads (0.00% of total) failing ReassignOneMappingQualityFilter
I also tried without "--filter_mismatching_base_and_quals" but it returned this ERROR message:
##### ERROR MESSAGE: SAM/BAM/CRAM file htsjdk.samtools.SamReader$PrimitiveSamReaderToSamReaderAdapter@36c1b6ff is malformed. Please see https://software.broadinstitute.org/gatk/documentation/article?id=1317for more information. Error details: BAM file has a read with mismatching number of bases and base qualities. Offender: 23570007 [51 bases] [0 quals]. You can use --defaultBaseQualities to assign a default base quality for all reads, but this can be dangerous in you don't know what you are doing.
I'm fairly new to GATK and can't really think of a reason why this would happen, so I would really appreciate if anyone can help.