ATAC-seq analysis of transgenic human cancer cell line expressing subunits of meiotic cohesin complexes (RAD21L mei-Cohesin)

The goal of the study was to investigate the epigenomic properties of somatic cells in response to the induction of mei-cohesin subunits using Tet-on inducible transgenes activated by an rtTA transgene and doxycicline. The study involved applying ATAC-seq (Assay for Transposase-Accessible Chromatin with high-throughput sequencing) analysis of stable transgenic human cancer cell line DLD-1, which expresed Tet-inducible combinations of mei-cohesin subunits. The ATAC-seq method enables mapping DNA accessibility in chromatin with hyperactive Tn5 transposase inserting specific adapters into genome. Overall design: The DLD-1 (ATCC® CCL-221™) based cell lines were first stably infected with rtTA, followed by Tet-On-STAG3, Tet-On-SMC1beta, and Tet-On RAD21L lentiviruses, to generate two multi-transgenic cell lines. The populations of transgenic cells selected for marker drug-resistance were cloned to generate single transgenic clones used for ATAC-seq experiments.

本研究旨在探究体细胞经rtTA转基因（rtTA transgene）与多西环素（doxycycline）激活的Tet-on诱导型转基因诱导减数分裂黏连蛋白亚基后的表观基因组特性。
本研究针对稳定表达Tet诱导型减数分裂黏连蛋白亚基组合的转基因人类癌细胞系DLD-1，开展了ATAC-seq（转座酶可及性高通量测序，Assay for Transposase-Accessible Chromatin with high-throughput sequencing）分析。ATAC-seq技术借助高活性Tn5转座酶（Tn5 transposase）将特异性接头插入基因组，可实现染色质内DNA可及性的图谱绘制。
整体实验设计如下：以DLD-1细胞（ATCC® CCL-221™）为基础细胞系，先稳定感染rtTA慢病毒，随后分别感染Tet-On-STAG3、Tet-On-SMC1beta及Tet-On RAD21L慢病毒，构建得到两种多转基因细胞系；通过标记性耐药筛选获得的转基因细胞群经克隆后，得到用于ATAC-seq实验的单转基因克隆。