The wucai genome and DNA methylation regulation on the inner leaves' yellowing response to low temperature.

Wucai (Brassica rapa), a widely cultivated non-heading Chinese cabbage in autumn and winter in China, has been limited in its genetic and genomic analyses due to the lack of a reference genome. Here, we have assembled a high-quality chromosome-level genome assembly of wucai, which is 480.57 Mb in length, with a scaffold N50 of 46.53 Mb and a contig N50 of 4.45 Mb. We annotated 42 634 protein-coding genes in the B. rapa W7-2 genome and 55.08% of repetitive sequences. Additionally, we performed DNA methylome analysis to investigate the characteristic yellowing response of wucai inner leaves to low temperature. CHH methylation levels increased under low-temperature conditions, while a slight decrease was observed under normal temperatures with more hypo-differentially methylated regions. The expression levels of DNA methyltransferases BrCMT2 and BrDRM2 were significantly up-regulated under low-temperature conditions but down-regulated at normal temperature. Furthermore, chlorophyll metabolism genes, BrHemA, BrHemL, BrHemD, BrCLH2, BrCHLP, BrSGR, and BrPPD, were identified as differentially expressed, which exhibited elevated CHH methylation levels in their promoter regions across three stages. Leaves of Nicotiana benthamiana transiently overexpressing BrCLH2.1 exhibited a slight degreening phenotype compared to wild type. The application of a DNA methylation inhibitor into the inner leaves of W7-2 induced obvious bleaching, and the total chlorophyll content decreased significantly. In conclusion, the genome data of W7-2 provide a valuable resource for functional gene research and genetic breeding of non-heading Chinese cabbage.