Using reduced representation bisulfite sequencing (RRBS) to characterize genomic DNA methylation

Background

• Seminal ​paper to introduce RRBS. The workflow below illustrates how to use ​bismark on the resources at the Whitehead Institute.

Step by step analysis

• QC
  • Use ​Trim Galore or ​another read trimmer to apply quality filters and remove adapters.
  • See our ​QC and preprocessing guidelines for details on running Trim Galore.
  • When . A command for Trim Galore paired end reads using gzipped fastq input can look like:

bsub trim_galore --paired --rrbs --fastqc -o trimmedReads /path/to/raw/data/reads_1.fq.gz /path/to/raw/data/reads_2.fq.gz

• Quantification of methylation calls
  • Bismark produces BAM file(s) of aligned reads and methylation calls.

bsub /path/to/bismark/bismark --genome /nfs/genomes/mouse_mm10_dec_11_no_random/bowtie/ -1 trimmedReads_1.fq.gz -2 trimmedReads_2.fq.gz

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• Extract methylation calls
  • After running bismark...
  • For paired ended data...:

bsub /path/to/bismark/bismark_methylation_extractor -p --gzip --bedGraph trimmedReads_bismark_bt2_pe.bam 

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