Question: what's wrong with htseq-count codes???
0
9 weeks ago
biologo • 20

hello, friends:

i was using the htseq-count to calculate the reads number, after run the tophat, then i isolate the uniqmapped reads, actually ,it was the paired-end reads, but i find uniqmapped reads only contain the single-end reads(no identical readsID),that is one thing that i fell confused??? next, i do like this:

samtools view -@ 5 -S uniqmap.sam -b -o uniqmap.bam
samtools sort -@ 5 -n uniqmap.bam -o sort.bam
samtools view -@ 5 -h sort.bam -o sort.sam
htseq-count -m union -s no sort.sam $GTF > union.out

and the result is nothing, err report that:

Warning: Read HWI-ST1434:279:HTFV7BCXX:2:1115:7046:27240 claims to have an aligned mate which could not be found. (Is the SAM file properly sorted?)
Warning: Read HWI-ST1434:279:HTFV7BCXX:1:1115:5605:41821 claims to have an aligned mate which could not be found. (Is the SAM file properly sorted?)
Warning: Read HWI-ST1434:279:HTFV7BCXX:1:2111:7809:19690 claims to have an aligned mate which could

can you help me figure it out???

RNA-Seq • 842 views
ADD COMMENTlink 2.7 years ago biologo • 20 • updated 9 weeks ago lieven.sterck 5.1k
1
2.7 years ago
WouterDeCoster 39k
Belgium

There is no need to filter the uniquely mapped reads yourself (likely the step which went wrong). Htseq count will be default only use read with mapQ >= 10.

ADD COMMENTlink 2.7 years ago WouterDeCoster 39k

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