#302 Bowtie2 alignment speed very slow for many Ns in the reference sequence

[Felix Krueger]

Lately I have been experiencing very slow mapping speeds with Bowtie 2 against a genome containing many ‘Ns’ and was wondering if anyone has experienced the same or know a solution to this.

I have generated some mouse strain genomes containing Ns at known SNP positions to the Black6 reference genome. Running standard 50bp paired-end alignments on 10 cores took more than 10 days to complete for ~200M sequence pairs, which doesn’t sound that it is meant like that. I have since tested a few things, such as the latest version (2.2.0) or the previous one (2.2.1) but that’s not the issue. Also reducing the --score-min parameter didn’t speed it up noticeably. I then took 1M test reads and aligned them to the following 3 genomes:

1) Genome containing 18M Ns, time: ~2h
2) Genome containing 4M Ns, time: 30 mins
3) Black6 reference genome, time : 2 mins

I have noticed that the 3rd index file increased in size from 5858 bytes for Black6 to 156842630 bytes for the N-strain. Is this the index file describing the position of Ns?
Do I just have to accept that Ns in the genome slow Bowtie 2 down >50-fold or is there any known cure for this?

[Ben Langmead]

Hi Felix,

A couple questions.

Are the Ns distributed fairly evenly across the genome?

Can we get a copy of the FASTA file you're using?

Best,
Ben

[Felix Krueger]

Hi Ben,

Thanks for getting back to me so quickly. Yes I suppose the Ns should be distributed evenly over all chromosomes but chrY. I have uploaded the FastA files as well as the *bt2 index files to the following server:

Connection Details

Hostname ftp2.babraham.ac.uk
Username ftpusr92
Password vZ4QtKE1
FTP URL ftp://ftpusr92:vZ4QtKE1@ftp2.babraham.ac.uk

I have run another test on 5M reads today and it took 9h 13 mins using 10 cores, which is pretty consistent with the 1M test sequences and close to 2h I had seen from a different file yesterday.
I already got a suggestion on SeqAnswers to use BBmap instead because it is much better and faster than every other tool on the planet, did you get a chance to look at these claims?

Again, many thanks,
Best,
Felix

---

From: Ben Langmead [ben_langmead@users.sf.net]
Sent: 26 February 2014 20:28
To: [bowtie-bio:bugs]
Subject: [bowtie-bio:bugs] #302 Bowtie2 alignment speed very slow for many Ns in the reference sequence

Hi Felix,

A couple questions.

Are the Ns distributed fairly evenly across the genome?

Can we get a copy of the FASTA file you're using?

Best,
Ben

---

[bugs:#302]http://sourceforge.net/p/bowtie-bio/bugs/302/ Bowtie2 alignment speed very slow for many Ns in the reference sequence

Status: open
Created: Wed Feb 26, 2014 01:50 PM UTC by Felix Krueger
Last Updated: Wed Feb 26, 2014 01:50 PM UTC
Owner: nobody

Lately I have been experiencing very slow mapping speeds with Bowtie 2 against a genome containing many ‘Ns’ and was wondering if anyone has experienced the same or know a solution to this.

I have generated some mouse strain genomes containing Ns at known SNP positions to the Black6 reference genome. Running standard 50bp paired-end alignments on 10 cores took more than 10 days to complete for ~200M sequence pairs, which doesn’t sound that it is meant like that. I have since tested a few things, such as the latest version (2.2.0) or the previous one (2.2.1) but that’s not the issue. Also reducing the --score-min parameter didn’t speed it up noticeably. I then took 1M test reads and aligned them to the following 3 genomes:

1) Genome containing 18M Ns, time: ~2h
2) Genome containing 4M Ns, time: 30 mins
3) Black6 reference genome, time : 2 mins

I have noticed that the 3rd index file increased in size from 5858 bytes for Black6 to 156842630 bytes for the N-strain. Is this the index file describing the position of Ns?
Do I just have to accept that Ns in the genome slow Bowtie 2 down >50-fold or is there any known cure for this?

---

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[Val]

Fixed for the next release.