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Old 06-21-2017, 10:20 AM   #1
rtleenay
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Location: North Carolina

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Default Low mapping rate for smallRNA Seq

Hello,

I am trying to align small/miRNAs to a fully assembled bacterial genome, and have been only getting 7% of reads to align. Of those 7%, almost all of them are only 10nt in length (or shorter). The size distribution has a peak around 30nt in length, and then slowly tails to 150. Most of the quality scores are 34 or higher.

Can anyone provide any suggestions? I have tried Bowtie2 default settings, local alignment, and endtoend with a 4nt seed. All have given me the same problem.

Thank you!
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Old 06-21-2017, 10:26 AM   #2
rtleenay
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It is important to note that we did not do rRNA depletion - it was not available for our bacterium. I have also tried both local and endtoend alignment with Bowtie2 and I continually get a low percentage. The samples were treated with a 5' de-phosphorylase to look for RNA processing, although that should not explain the extreme lack of mapping %.
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Old 06-21-2017, 10:47 AM   #3
GenoMax
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You should try a non-gapped aligner like bowtie v.1 for this. Be sure to remove/trim any kit specific adapters/illumina adapters before alignment.
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