DNA甲基化
亚硫酸氢盐测序
生物
拟南芥
表观遗传学
表观遗传学
亚硫酸氢盐
转座因子
DNA
基因组
胚胎
甲基化DNA免疫沉淀
计算生物学
遗传学
基因
细胞生物学
基因表达
突变体
作者
Hyunjin Yoo,Kyunghyuk Park,Jae‐Hoon Lee,Seunga Lee,Yeonhee Choi
出处
期刊:Molecules and Cells
[Korean Society for Molecular and Cellular Biology]
日期:2021-08-01
卷期号:44 (8): 602-612
被引量:3
标识
DOI:10.14348/molcells.2021.0084
摘要
DNA methylation is an important epigenetic mechanism affecting genome structure, gene regulation, and the silencing of transposable elements.Cell-and tissuespecific methylation patterns are critical for differentiation and development in eukaryotes.Dynamic spatiotemporal methylation data in these cells or tissues is, therefore, of great interest.However, the construction of bisulfite sequencing libraries can be challenging if the starting material is limited or the genome size is small, such as in Arabidopsis.Here, we describe detailed methods for the purification of Arabidopsis embryos at all stages, and the construction of comprehensive bisulfite libraries from small quantities of input.We constructed bisulfite libraries by releasing embryos from intact seeds, using a different approach for each developmental stage, and manually picking single-embryo with microcapillaries.From these libraries, reliable Arabidopsis methylome data were collected allowing, on average, 11fold coverage of the genome using as few as five globular, heart, and torpedo embryos as raw input material without the need for DNA purification step.On the other hand, purified DNA from as few as eight bending torpedo embryos or a single mature embryo is sufficient for library construction when RNase A is treated before DNA extraction.This method can be broadly applied to cells from different tissues or cells from other model organisms.Methylome construction can be achieved using a minimal amount of input material using our method; thereby, it has the potential to increase our understanding of dynamic spatiotemporal methylation patterns in model organisms.
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