Spermatozoa RRBS data: Post-Bismark aligned files extracted for the methylation call for every single C in the CpG context of sperm cells of Murrah buffalo bulls under heat stress

The DNA was isolated from purified spermatozoa samples obtained during hot summer season from the Murrah buffalo bulls (n=10) and were sequenced using RRBS. The animals belonged to two groups, seasonally affected (SA=5) and seasonally not affected (SNA=5), based on the semen quality parameters (sperm viability, hypo-osmotic swelling test, and acrosomal integrity), which significantly varied across the four seasons between the two groups. The study is the first of its kind to generate the methylome of heat stress affected Murrah buffalo bulls from the sperm cells. This data potentially paves the path to understanding the epigenetic regulation of heat stress on altered sperm function and semen quality.

Methods
The sequences were aligned against the Bubalus bubalis assembly using the Bismark suite.The unsorted BAM files were processed for the next step, where the information regarding the methylated CpGs was extracted. This particular step is not mandatory to be performed, but generating the methylation context-wise report can assure an error-free DMC calculation in further steps. The methylation base extraction is enabled by the script within the Bismark suite 'bismark_methylation_extractor'. Depending on the context (CpG, CHG, CHH), the positions of every cytosine in the BAM file were documented to a new output file, with methylated cytosines labelled as forward reads and non-methylated cytosines labelled as reverse reads.