Can anyone suggest resources for troubleshooting why my alignment using Bowtie1.1.1 is incorrect?
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2 days ago
nessj • 0

My bowtie command :

bowtie GCF_000001215.4_Release_6_plus_ISO1_MT_genomic -q SRR8191524.fastq -v 2 -m 1 -3 1 -S 2> ./SRR8191524.out > ./SRR8191524.sam


Samtools flagstat output of bam file:

8639841 + 0 in total (QC-passed reads + QC-failed reads)
0 + 0 secondary/ 0 + 0 supplementary / 0 + 0 duplicates
5640087 + 0 mapped (65.28% : N/A)
0 + 0 paired in sequencing/ 0 + 0 read1/ 0 + 0 read2/ 0 + 0 properly paired (N/A : N/A)/ 0 + 0 with itself and mate mapped/ 0 + 0 singletons (N/A : N/A)/ 0 + 0 with mate mapped to a different chr/ 0 + 0 with mate mapped to a different chr (mapQ>=5)

flagstat Bowtie1.1.1 samtools • 134 views
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Looks like 65% of your reads have mapped. Depending on the dataset this is not a bad result. Why do you think this alignment is incorrect?

bowtie v.1.x does only ungapped alignments so if you expect there to be splicing then it would not be the right aligner to use.

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thank you for your help! i am certainly new to this data processing --and so I thought there would be statistics in the alignment, but everything fills in at 0 (0 paired in sequencing, 0 properly paired, etc)

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Since you have a single-end dataset many other stats that deal with paired-end data are 0.

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**revise: I appreciate your help GenoMax --i think i can get around the below issue now that I know it isn't my alignment!

also my MACS2 callpeak output error is : "Too few paired peaks (0) so I can not build the model! Broader your MFOLD range parameter may erase this error. "