Bisulfite-Seq

Scripts related to processing of bisulfite sequencing data via Snakemake and the SLURM workload manager for parallel computing.

Makes use of methylKit for differential analysis.

Makes use of conda environment to provide relevant software.

Standard Steps include:

1. Check quality of raw data with FastQC
2. Trim with TrimGalore (-rrbs option for RRBS data)
3. Check quality of trimmed reads with FastQC
4. Align trimmed reads with Bismark
5. Collect trimming and alignment statistics into tab delimited text files
6. Extract methylation data with bismark_methylation_extractor
7. Produce canonical coverage files for downstream use
8. Perform initial data exploration, including:

• number of CpGs covered per sample at various coverage cutoffs
• "coverage plots" from methylKit
• "methylation stats plots" from methylKit
• dimension reduction and visualization of samples, including PCA and Nonnegative Matrix Factorization (NMF) + tSNE

9. Perform DMR analysis via methylKit to find differentially methylated regions between two groups
10. Produce a DMR bed file track for a genome browser
11. Produce 3 bed files per comparison for use with GREAT pathway analysis

• Both hyper and hypomethylated regions
• Hypermethylated regions only
• Hypomethylated regions only