DNA Adenine Methyltransferase Identification (DamID) Coupled to Next Generation Sequencing For Profiling Lamina Associated Domains

Purpose: To compare the interaction of various proteins (in particular Lap2b and Lamin A) at the nuclear periphery with the Lamina Associated Domains (LADs) Results: Using an optimized data analysis workflow (LADetector, available at https://github.com/thereddylab/LADetector), We found near identical sequences within the genome-wide maps indicating that LAP2ß and Lamin A, and hence LaminB1, are in close proximity to all LADs, suggesting that these proteins are not involved in discriminating subsets of LADs. Overall design: LAD profiles, of MEFs, corresponding to various proteins at the nuclear periphery (LMNA, Lap2b) were generated following the DamID protocol (L.Guelen et al, Nature 2008), library randomization and sequencing preparation using Illumina Paired End Sequencing Kit and sequenced on the Illumina HiSeq 2000.

研究目的：比较核周各类蛋白质（尤其是Lap2b与核纤层蛋白A（Lamin A））与核纤层结合结构域（Lamina Associated Domains, LADs）的相互作用。
研究结果：采用优化的数据分析流程（LADetector，开源地址：https://github.com/thereddylab/LADetector），我们在全基因组图谱中检测到近乎完全一致的序列信号，结果显示LAP2β与核纤层蛋白A（Lamin A）以及核纤层蛋白B1（LaminB1）均与所有LADs紧密毗邻，提示上述蛋白质并未参与区分不同子集的LADs。
实验整体设计：按照DamID技术流程（L.Guelen等，《Nature》，2008年），为小鼠胚胎成纤维细胞（Mouse Embryo Fibroblasts, MEFs）的核周各类蛋白质（LMNA、Lap2b）制备LAD图谱；通过Illumina Paired End Sequencing Kit完成文库随机化与测序文库构建，并使用Illumina HiSeq 2000测序平台完成测序。