BISULFITE-SEQ ANALYSIS PIPELINE USING SNAKEMAKE AND BISMARK

SUMMARY This Snakemake pipeline automates the processing of paired-end bisulfite sequencing data for methylation analysis in Nasonia vitripennis. It performs adapter trimming, genome preparation, read alignment (both experimental and lambda controls), methylation extraction, and final CpG merging. CONFIGURATION Sample names must be defined in an external `config.yaml` file: config.yaml: samples: - sample1 - sample2 ... KEY STEPS 1. Trimming: Adapter trimming and quality filtering using `cutadapt` 2. Genome Preparation: Bismark-compatible indexing of both experimental and lambda genomes 3. Alignment: Paired-end alignment using `bismark` with Bowtie2 4. Methylation Extraction: Extract methylation calls from BAM files 5. CpG Merging: Collapse strand-specific CpG methylation into unified calls RULES OVERVIEW - `all`: Defines final output files to build for all samples - `trimming`: Trims adapters from raw fastq files - `genome_prep`: Prepares the experimental genome for Bismark - `lambda_genome_prep`: Prepares the lambda genome for control alignment - `alignment`: Aligns reads to the experimental genome - `lambda_alignment`: Aligns reads to the lambda genome (conversion rate check) - `meth_extraction`: Extracts methylation data from aligned BAM - `merge_cpgs`: Merges complementary CpG calls into a single output file REQUIRED MODULES (LOADED IN SHELL COMMANDS) - `py-cutadapt/4.4-jv44fem` - `bowtie2/2.5.1-s4zfmrn`