Samtools Flagstat Output

Question

[Nitram]

For past two hours I tried to browse around for samtools flagstat output information, unfortunately could not find much related to my problem - this is my samtool flagstat output for single end reads - does anyone think that the tophat alignment did not work well. Please let me know. Any possible replies would be greatly appreciated. Thanks in advance.

$ samtools flagstat accepted_hits.bam

18109074 + 0 in total (QC-passed reads + QC-failed reads) 0 + 0 duplicates 18109074 + 0 mapped (100.00%:nan%) 0 + 0 paired in sequencing 0 + 0 read1 0 + 0 read2 0 + 0 properly paired (nan%:nan%) 0 + 0 with itself and mate mapped 0 + 0 singletons (nan%:nan%) 0 + 0 with mate mapped to a different chr 0 + 0 with mate mapped to a different chr (mapQ>=5)

Answer 1

[Wen.Huang]

your output actually says all were mapped. I am pretty sure that top hat only outputs mapped reads. you need to compare the number to the input file.

[Wen.Huang]

It has to be multi-reads.

try

samtools view accepted_hits.bam | cut -f1 | sort | uniq | wc -l

this should give you the count of unique reads

[Wen.Huang]

It has to be multi-reads. try "samtools view accepted_hits.bam | cut -f1 | sort | uniq | wc -l" this should give you the count of unique reads

[Nitram]

Thanks so much for your reply. My input fastq file has only 17,264,285 reads, then how could the number of mapped reads (~18,109,074) be higher.

[Swbarnes2]

Is it possible that bowtie is outputting repetative reads multiple times? samtools flagstat would not try to account for this.

[Nitram]

Thanks so much. It helped !