Bowtie2 is probably the most widely used aligner because of it's speed. Burrow-wheeler (BW) algorithms (including bwa) tend to be faster. However, they have limitations when it comes to aligning very short reads (e.g. gRNA). Also, setting maximum number of mismatches allowed is complicated by the seed length, overlaps and other parameters.

I wonder if there is any better multi-purpose aligner out there. May be with algorithm other that BW. One which allows special cases e.g. allowing shord reads and high number of mismatches.

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    $\begingroup$ How long is "very short"? Bowtie2 is not optimized for <=36bp query sequences; nor most aligners developed in the past 5 years or so. Bowtie1 or bwa-aln will work better. $\endgroup$
    – user172818
    Jun 18, 2018 at 1:53
  • $\begingroup$ @user172818 When you say "not optimized", do you mean using the default settings, or more generally ? $\endgroup$
    – bli
    Jun 18, 2018 at 9:52
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    $\begingroup$ @bli in general. Aligning 100bp reads is a very different problem from aligning <36bp reads. $\endgroup$
    – user172818
    Jun 18, 2018 at 12:05
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    $\begingroup$ If I have learned one thing working with bioinformatics over the years, that is there is no such thing as a "multi-purpose" or "overall best" tool out there. Different algorithms are good for different types of data. My recommendation would be to arrange your workflow in a modular way and plug in the best aligner for the type of data you are working on, rather than relying on a "best" solution. $\endgroup$
    – posdef
    Feb 4, 2019 at 12:00
  • $\begingroup$ As of 2020-09-09, according to google scholar, the 2012 bowtie2 paper has 21647 citations, the 2009 bwa paper has 25045 citations. It may be true that bowtie2 is the most widely used aligner, but not probable. $\endgroup$
    – winni2k
    Sep 9, 2020 at 12:17

1 Answer 1


Bowtie2 is no longer the fastest aligner. Salmon and Kallisto are much faster, but have been designed to optimise RNASeq mapping. Their speed is gained from avoiding a strict base-to-base alignment, but they can output mostly-aligned reads (i.e. position-only, without local alignment) as pseudo-alignments. See here for more details.

Both Kallisto and Salmon can do additional bootstrapping that is interpreted by sleuth for improved performance in isoform detection. They can also output counts that are equivalent to read-level counts from other programs, which can then be used by other downstream gene-based differential expression analysis software (e.g. DESeq2). Salmon has additional options that can correct mappings for sequence-level and GC bias.

HISAT2 is from the same group as Bowtie2, and does the same sort of stuff, but with a few optimisations added on top. In particular, it's much better at working out split reads from RNASeq runs. Like Bowtie2, it will do local alignment of reads.

For quick alignment of long reads, minimap2 works well. For high-accuracy alignment (but comparatively slower), LAST works well.

Most bioinformaticians seem to prefer STAR for things that Bowtie2 was previously used for. I'm not yet convinced it's a better alternative, and currently prefer HISAT2 for high accuracy short-read alignment.

  • $\begingroup$ Seconded for kallisto, it is very good. But currently downstream support is limited to the other tool from the pachter lab for differential gene expression analysis, called sleuth. $\endgroup$
    – NatWH
    Jun 18, 2018 at 0:45
  • $\begingroup$ One advantage to STAR aligner in Bioinformatics and the reason it is used at my work, is the possibility of reading Chimeric alignments (for detecting e.g. circular RNA through custom coding). $\endgroup$ Jun 18, 2018 at 9:58
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    $\begingroup$ Salmon and Kallisto aren't really aligners and are only useful for RNA-seq, though they are great and very quick. @NatWH Kallisto can also be used with DESeq2 or edgeR. $\endgroup$ Jun 19, 2018 at 20:14
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    $\begingroup$ @Jared Andrews alright. I'll leave my comments up in case any future visitors benefit from the links which have been posted. $\endgroup$
    – NatWH
    Jun 20, 2018 at 22:23
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    $\begingroup$ Kallisto can output position. What it doesn't do is a careful local alignment (and the question didn't specify whether local alignment was important). I indicated other alternatives where local alignment is important. $\endgroup$
    – gringer
    Feb 1, 2019 at 20:04

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