DNA methylation analysis for mouse embryonic stem cells deficient for Smad1 and Smad5 or for Bmp activated subpopulations.

In this study we determine the profile of DNA methylation by RRBseq of mESC in the absence of Smad1 and Smad5 and in subpopulation of mESC with different levels of BMP-SMAD activation. We observed a general loss of DNA methylation associated with low or absent BMP-SMAD signalling in mESCs. Overall design: DNA methylation analysis using RRBS was performed on 3 biological replicates of BRE negative and positive mESC subpopulations, which were collected in pairs at 3 different times. DNA methylation analysis using RRBS was performed on Smad1/5 floxed (FL) and knockout (KO) mESC. Two different parental cell lines were used. For each parental cell line we analyzed one Smad1/5 FL sample and two Smad1/5 KO samples, resulting in respectively two and four biological replicates for the FL and KO conditions.

本研究通过RRBS（Reduced Representation Bisulfite Sequencing，简化亚硫酸氢盐测序）分析了缺失Smad1与Smad5的小鼠胚胎干细胞（mouse embryonic stem cells, mESC），以及不同BMP-SMAD激活水平的小鼠胚胎干细胞亚群的DNA甲基化谱。研究团队观察到，在小鼠胚胎干细胞中，BMP-SMAD信号通路活性低下或缺失时，会伴随DNA甲基化的普遍丢失。
实验整体设计：针对BRE阴性与阳性的小鼠胚胎干细胞亚群，分别设置3组生物学重复样本，所有样本均于3个不同时间点成对收集，并通过RRBS开展DNA甲基化分析。针对Smad1/5 floxed（FL）与敲除（KO）的小鼠胚胎干细胞，同样通过RRBS开展DNA甲基化分析。本实验使用了两种不同的亲本细胞系，针对每一种亲本细胞系，分别分析1个Smad1/5 FL样本与2个Smad1/5 KO样本，最终FL组与KO组的生物学重复样本数分别为2与4。