Abstract:
Cleavage Under Targets and Tagmentation (CUT&Tag) provides high-resolution sequencing libraries for profiling diverse chromatin components. This protocol details the steps to generate CUT&Tag libraries from fresh or frozen tissues. This CUT&Tag workflow has nine main steps: isolation of nuclei from tissues, binding of nuclei to Concanavalin A-coated beads, binding of the primary antibody, binding of the secondary antibody, binding pA-Tn5 adapter complex, tagmentation, DNA extraction, PCR, and post-PCR cleanup and size selection. This protocol enabled us to generate and sequence CUT&Tag libraries across a broad range of fresh and frozen tissue types.
摘要:
靶标下的切割和标签(CUT&Tag)提供高分辨率测序文库,用于分析不同的染色质组分。该协议详细说明了从新鲜或冷冻组织生成CUT&标签库的步骤。这个CUT&标记工作流程有九个主要步骤:从组织中分离细胞核,细胞核与刀豆蛋白A包被的珠子结合,一级抗体的结合,第二抗体的结合,结合pA-Tn5衔接子复合物,标签化,DNA提取,PCR,以及PCR后的清理和大小选择。该协议使我们能够在广泛的新鲜和冷冻组织类型中生成和测序CUT&标签库。