Question: Read alignment using Bowtie2
gravatar for Ina
10 months ago by
Ina0 wrote:

Hello everyone!

I am in need of using Bowtie2 to find all possible alignments for a set of reads (each 23bp long) to the human genome. The maximum number of mismatches that can be tolerated is 6. But since Bowtie2 allows mismatches only upto 1, I had to set the mismatch penalty for alignment score accordingly to get the result. Since my laptop does not have the capacity to perform this operation, I am searching for interfaces that provide access to Bowtie2. I found Galaxy to be useful but unfortunately, I cannot see an option to set the alignment score in it. Does anyone know of any other tool that provides an option for Bowtie2 with alignment score?


score alignment bowtie2 • 421 views
ADD COMMENTlink written 10 months ago by Ina0

For ultra-short reads it is recommended to use bowtie (aka bowtie1) which allows you to explicitely set the number of tolerated mismatches. From the bowtie2 FAQs:

For reads longer than about 50 bp Bowtie 2 is generally faster, more sensitive, and uses less memory than Bowtie 1. For relatively short reads (e.g. less than 50 bp) Bowtie 1 is sometimes faster and/or more sensitive.

Be aware that with this short length you'll get plenty of multimappers especially if allowing that many mismatches which is 26% of the total read length. What experiment is this, maybe there are better alternatives?

ADD REPLYlink modified 10 months ago • written 10 months ago by ATpoint36k


Thank you ATpoint for your suggestion.

So this is related to CRISPR-CAS9. I am working with off-target predictions for my thesis and was looking at all scientific papers related to CRISPR. I found one and decided to use their datasets. The datasets were not available directly and the method used to obtain them was specified which I had mentioned below:

They have around 30 sgRNAs which are each 23bp long and are aligning the sgRNAs to human reference genome with a tolerance of upto 6 mismatches using Bowtie2.

In order to access the exact dataset, I thought I need to replicate the method they used. To do so, I collected the sgRNAs from their paper and tried to use Bowtie2 to align them against human genome with upto 6 mismatches. I decided that only Bowtie2 can be used since the output from different tools may not be deterministic.

ADD REPLYlink modified 10 months ago • written 10 months ago by Ina0
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