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Bismark specifications


Unique identifier OMICS_00575
Name Bismark
Software type Package/Module
Interface Command line interface
Restrictions to use None
Input format FASTQ
Output format SAM, tab-delimited text
Operating system Unix/Linux
Programming languages Perl
License GNU General Public License version 2.0
Computer skills Advanced
Version 0.14.3
Stability Stable
Maintained Yes




No version available


Publication for Bismark

Bismark citations


B cell activation and plasma cell differentiation are inhibited by de novo DNA methylation

Nat Commun
PMCID: 5953949
PMID: 29765016
DOI: 10.1038/s41467-018-04234-4

[…] RRBS FASTQ files were quality trimmed using FastQC (v0.11.5) and mapped to the mouse genome (mm9) using Bismark (v0.16.3) using the following options: “--bam --chunkmbs 1024 –multicore 8”. Mapped BAM files were sorted with SAMtools (v0.1.19-96b5f2294a) and methylation calls were extracted using the “Rsa […]


Genome wide profiling of long non coding RNAs from tomato and a comparison with mRNAs associated with the regulation of fruit ripening

BMC Plant Biol
PMCID: 5935960
PMID: 29728060
DOI: 10.1186/s12870-018-1300-y

[…] nome bisulfite sequencing data sets derived from 39 dpa and 52 dpa fruit were retrieved from the SRA database (SRA046092). Raw sequence data were mapped to the tomato genome assembly build 2.50 using Bismark 0.16.3 [] with the Bowtie2 (version 2.2.8) alignment tool []. The percent methylation of cytosines in a CpG, CHG or CHH context was calculated by the proportion of 100* (methylated reads)/ (me […]


Clinical Remission of Sight Threatening Non Infectious Uveitis Is Characterized by an Upregulation of Peripheral T Regulatory Cell Polarized Towards T bet and TIGIT

Front Immunol
PMCID: 5943505
PMID: 29774027
DOI: 10.3389/fimmu.2018.00907

[…] te the molarity of the library before sequencing. 150-bp Paired-end sequencing was performed using the Miseq Illumina sequencer (Illumina, Inc.) to a total of 400,000 reads passing filter per sample. Bismark () was used to align bisulfite-treated reads to a human reference genome and calculate the cytosine methylation calls at the target sites. Biostatistical analysis generated beta (%) methylatio […]


Long ncRNA A ROD activates its target gene DKK1 at its release from chromatin

Nat Commun
PMCID: 5915440
PMID: 29691407
DOI: 10.1038/s41467-018-04100-3

[…] sequencing in MCF-7 cells was done using the SureSelect Methyl-Seq Target Enrichment System from Agilent. Data analysis (including read mapping, deduplication and methylation calling) was done using Bismark. Data available under GEO (NCBI) accession number GSE69505. […]


Association and cis mQTL analysis of variants in CHRNA3 A5, CHRNA7, CHRNB2, and CHRNB4 in relation to nicotine dependence in a Chinese Han population

PMCID: 5904126
PMID: 29666375
DOI: 10.1038/s41398-018-0130-x

[…] as identified using the Illumina HiSeq X Ten platform with an average of about 700 million (±75 million) 150-bp paired-end reads per sample. Clean reads were mapped to the hg19 reference genome using Bismark. We first combined two strands of information of CpG sites and then excluded those CpGs with <5 reads or that overlapped with common variants in the Chinese Han genome (MAF > 0.05). The MAF of […]


Data quality of whole genome bisulfite sequencing on Illumina platforms

PLoS One
PMCID: 5905984
PMID: 29668744
DOI: 10.1371/journal.pone.0195972
call_split See protocol

[…] nd of R1 to remove bases derived from the sequence tag introduced in the library preparation procedure. Alignment to the human reference assembly GRCh37 and methylation calling was performed with the Bismark software [] and the pipeline tool ClusterFlow []. For TSDM libraries the initial 6 base pairs of each reads were ignored in the methylation calling procedure, to avoid random priming biases. G […]


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Bismark institution(s)
Bioinformatics Group, The Babraham Institute, Cambridge, UK
Bismark funding source(s)
Biotechnology and Biological Sciences Research Council (BBSRC)

Bismark reviews

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Miklós Laczik

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Bismark is the definitive tool for (primary and secondary) bisulfite sequencing analysis. It is easy to use, very stable, well maintained, I had zero issues with it over the years. Not only it aligns the reads to a reference genome, it's methylation extractor module produces various useful outputs in standard formats (sam, bam, bedGraph etc.) which are customizable too (e.g. separate results by strand, show only CpGs or CHG, CHH contexts separately, show data on all Cs on a genome or only the covered ones, produce UCSC compatible bedGraph files with 0-based half-open coordinates or alternative tab delimited files with 1-based coordinates, it can remove spaces from read names etc.); it really opens the door for all types of downstream analyses. The documentation is also superb, very well written with clear examples and a complete guide with all options, it leaves nothing to guess. There are even specific analysis recommendations for different applications and commercially available kits (e.g. when to remove duplicates, when to expect a non-directional library etc.). I'm generally a big fan of Babraham tools, Iáve been using Bismark for at least 5 years for RRBS and WGBS, it's still my number one choice and it gets better and better.
If I really want to nitpick and find a negative point, I would say the parallelization is a bit of a hassle if you have limited resources, it's not as straightforward as setting the number of threads to use, you need some experimenting with parameters like --multicore and -p to find out how many actual cores (and RAM) are used for a typical run with your settings. Also, itás not the fastest tool in the world, for example BSMAP is said to be faster (especially in RRBS mode), but then again in side-by-side comparisons I always managed to map more reads with Bismark, and I think the speed is quite reasonable, I'm happy with it.

Overall Bismark is a wonderful tool, I can highly recommend it for any (RR)BS data analysis!
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Anonymous user #5076

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Bismark is a great tool for the alignment of BS-seq reads.