Protocol for low-input whole-genome bisulfite sequencing in Arabidopsis

DNA methylation profiling of rare plant samples is limited by the scarcity of input DNA. Here, we present a protocol for low-input whole-genome bisulfite sequencing (WGBS) in Arabidopsis in which DNA inputs as low as 50 pg can generate methylomes comparable to conventional WGBS. We describe steps for tissue cryomilling, Proteinase K lysis, and direct bisulfite conversion. We then detail procedures for post-bisulfite adapter tagging library construction and MethylStar-based alignment and methylation calling.