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Creating reference Index using Bowtie2

Hi all,

I'm trying to create reference index for all human chromosomes 22+X+Y+M using bowtie2, but its getting stuck after performing some steps.

genome alignment next-gen assembly
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updated by Ram Ram • written by cvu cvu

You should add other details in order for us to help you. You should tell us what error it is throwing?

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written by Ashutosh Pandey Ashutosh Pandey

It's not giving any error, it is getting stuck. I left it running for 24hours on 8GB RAM system, but it stuck.

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updated by Ram Ram • written by cvu cvu

There are some per-built indexes available ftp://ftp.ccb.jhu.edu/pub/data/bowtie_indexes/, but it seems they are for Bowtie not for Bowtie2 as they have extension .ebwt.

So can I use these indexes for Bowtie2 also?

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updated by Ram Ram • written by cvu cvu

No, bowtie1 indices won't work for bowtie2.

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written by Devon Ryan Devon Ryan

Hi, Did you find the bug?

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updated by Ram Ram • written by SP SP

1 answer

Just download the prebuilt ones from iGenomes. Note that they don't seem to have the most recent human reference (GRCh38). If you really do need that, just stop the current bowtie2-build process and start it again, this is presumably a one-off even.

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written by Devon Ryan Devon Ryan

ya i want GRch38 reference index. 8GB RAM is enough or it needs more RAM?

i already tried it 3 times but it is not working.

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written by cvu cvu

I would expect that that's enough, but I don't have anything with that little RAM with which to test.

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written by Devon Ryan Devon Ryan

bowtie2-build -f GRch38.fasta ref_index

please check, above command line arguments i am using. please let me know, if its okay or i am missing something ?

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written by cvu cvu

You don't need -f, but that shouldn't be causing the problem. The command looks fine.

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written by Devon Ryan Devon Ryan

Do you want the UCSC or Ensembl chromosome names? I can just make the indices and put them on google drive or something for you.

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written by Devon Ryan Devon Ryan

I want Ensembl chromosome names (GRch38). please do me a favor, if you can.

Thanks a lot Devon Ryan!!

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updated by Ram Ram • written by cvu cvu

Can do. Is the "primary assembly" OK or do you want "top level" (i.e., with all of the patches and alternate haplotypes)?

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written by Devon Ryan Devon Ryan

basically, i'm gonna use this index for mapping reads to find SNPs and INDELs. and i've downloaded dbSNP human build 141(vcf file) based on GRch38 assembly.

please give me whichever is better with above mentioned dbSNP version.

i've also changed sequence identifiers in fasta files according to the name given in dbSNP.

so i can provide you the fasta sequences.

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written by cvu cvu

As mentioned below, a GRCh38 reference hasn't been made by Ensembl yet, so I can only offer the UCSC version.

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written by Devon Ryan Devon Ryan

Thanks a lot !!

Please let me know, which chromosomes you have included in index?

All 22+X+Y+M?

Also tell me the version, hg38?

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updated by Ram Ram • written by cvu cvu

GRCh38, which for UCSC includes all of the chromosomes and unplaced contigs and I believe alternate alleles as well.

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updated by Ram Ram • written by Devon Ryan Devon Ryan

BTW, that requires ~6 gigs of RAM to index.

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updated by Ram Ram • written by Devon Ryan Devon Ryan

I tried it with 8GB RAM. don't know what was the problem

Thanks a lot again. I've downloaded files.

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updated by Ram Ram • written by cvu cvu

Actually, Ensembl hasn't released a version for GRCh38 yet (it'll be in the next release, number 76).

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written by Devon Ryan Devon Ryan

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