Originally posted by TEFA
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Originally posted by EGrassi View PostWoa, I used samtools -n and htseq-count gave me all counts of zero...that's strange, I will check with IGV or similar tools but cuffdiff gave me fpkg values with the same gtf and bam file...
Simon
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Originally posted by Simon Anders View PostWas this by any chance a TopHat output? Somebody alerted me recently to a bug that causes mishandling of SAM lines that contain the "NH" flag, and that seems to appear in newer tophat output. I'll try to fix this ASAP.
Thank you,
E.
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Ok, on the whole data it falls back to all 0:
samtools view S1.sorted.bam.bam | sed 's/NM:i:\d//g' | htseq-count --stranded=no - /rogue/bioinfotree/prj/ewing-rnaseq/local/share/data/Homo_sapiens/UCSC/hg19/Annotation/Genes/genes.gtf > counts_S1_htseqq
I'm really a little surprised by the amount of errors and other glitches in rna seq software (this is a generic rant, not focused on htseq!)...it's a really complicated and new area, that's true, but right now using any of the existing tool seems a bet and to tell the truth I never feel confident on the results, as long as with every version results changes, and when something runs without errors I never know if that means that it worked or that an exception just had not screamed enough :/
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Originally posted by Simon Anders View PostSorry, it seems we made some mix-up between version 0.5.3p4 and 0.5.3p5. Essentially, p5 undid some fixes in p4, including the one for "*" qualities. Now, there is version 0.5.3p6, which should clean up this mess. Please let me know if you still have problems.
The error which I'm trying to solve is;
Error occured when processing SAM input (line 2305 of file HBI_10.sam): ("'seq' and 'qualstr' do not have the same length.", 'line 2305 of file HBI_10.sam') [Exception type: ValueError, raised in _HTSeq.pyx:808]
Has anyone of you come across this error? What might be the solution?
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Originally posted by maubp View PostSushant - what does line 2305 of your SAM file look like? Unless the quality entry is the special value '*' only then it is a likely an unrelated issue.
However, I converted my fastq files to fasta and then performed the alignment, which solved this problem.
But the mapping percentage I'm getting with SHRiMP is too low(2.77%), for the mirna aligning to mature.
What parameters should I consider?
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