Something screwing up SAM to BAM

[JackieBadger]

Hi,
I am having issues with converting my SAM to BAM.

I align my single end reads to a genomic scaffold in bowtie
#######
bowtie2 -f --very-sensitive-local -x Species_Index -U SingleEnds.fasta -S Alignment.sam --sam-rg-id "Sample_ID" --no-unal
#######

Alignment summary:

29835 reads; of these:
29835 (100.00%) were unpaired; of these:
121 (0.41%) aligned 0 times
2414 (8.09%) aligned exactly 1 time
27300 (91.50%) aligned >1 times
99.59% overall alignment rate

My SAM file has header @SQ lines denoting my reference contigs so I then convert to BAM using;
#######
samtools view -bS ALignment.sam > ALignment.bam
#######

I then check the BAM basic statistics:
######
samtools flagstat LS01ALignment.bam
######

Which gives a contradictory result to the initial SAM alignment summary:

29714 + 0 in total (QC-passed reads + QC-failed reads)
0 + 0 duplicates
29714 + 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)



Can anyone offer any pointers?

[swbarnes2]

Nothing's wrong. bowtie know how many reads there were total, and how many of them mapped, and how many didn't, but it isn't putting the unmapped reads in the .sam. That's what that --no-unal means, right?

[JackieBadger]

Ahhh frig I totally misinterpreted the BAM stats. Thanks for pointing that out..it's been a long day