染色质
生物
核糖核酸
分子生物学
组蛋白
染色质免疫沉淀
染色质重塑
细胞生物学
计算生物学
遗传学
基因
基因表达
发起人
作者
Nadiya Khyzha,Steven Henikoff,Kami Ahmad
标识
DOI:10.1038/s41592-022-01618-9
摘要
Abstract Whereas techniques to map chromatin-bound proteins are well developed, mapping chromatin-associated RNAs remains a challenge. Here, we describe Reverse Transcribe and Tagment (RT&Tag), in which RNAs associated with a chromatin epitope are targeted by an antibody followed by a protein A-Tn5 transposome. Localized reverse transcription generates RNA/cDNA hybrids that are subsequently tagmented by Tn5 transposases for downstream sequencing. We demonstrate the utility of RT&Tag in Drosophila cells for capturing the noncoding RNA roX2 with the dosage compensation complex and maturing transcripts associated with silencing histone modifications. We also show that RT&Tag can detect N6-methyladenosine-modified mRNAs, and show that genes producing methylated transcripts are characterized by extensive promoter pausing of RNA polymerase II. The high efficiency of in situ antibody tethering and tagmentation makes RT&Tag especially suitable for rapid low-cost profiling of chromatin-associated RNAs.
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