A combination of transcription factors mediates inducible interchromosomal contacts

Development of MAP-C (mutation analysis in pools by chromosome conformation capture), which involves performing 3C on a pool of mutants (generated by programmed oligonucleotide pools, error-prone PCR, or existing mutant collections), followed by amplification of 3C DNA using primers specific to a chromosomal contact of interest (with the mutagenized or barcode region included), and as a control, amplifying the mutagenized or barcode region regardless of ligation. The 3C and genomic libraries are amplified with primers that add sequencing and flowcell adapters, and then deep sequenced to quantify the abundance of each mutant/barcode. The ratio of abundance in the 3C compared to the genomic library reflects the extent to which each sequence variant participates in the chromosomal contact of interest.基于染色体构象捕获的混合池突变分析（MAP-C, mutation analysis in pools by chromosome conformation capture）的开发流程如下：首先对由程序化寡核苷酸池、易错PCR或现有突变体库构建的突变体混合池实施染色体构象捕获（Chromosome Conformation Capture，3C）实验；随后使用针对目标染色体相互作用位点且包含突变区域或条形码区域的特异性引物扩增3C DNA；同时设置对照实验：不考虑连接状态，直接扩增突变区域或条形码区域。随后使用带有测序接头与流动池接头的引物分别扩增3C文库与基因组文库，再通过深度测序定量各突变体/条形码的丰度。3C文库与基因组文库的丰度比值，可反映各序列变体参与目标染色体相互作用的程度。