RNA sequencing by direct tagmentation of RNA/DNA hybrids
RNA sequencing by direct tagmentation of RNA/DNA hybrids
复制标题
DOI：
10.1073/pnas.1919800117
复制
发表时间：
2020-02-11
影响因子：
11.1
通讯作者：
Wang, Jianbin
中科院分区：
文献类型：
--
作者：
Di, Lin;Fu, Yusi;Wang, Jianbin
通过RNA测序（RNA-SEQ）进行了转录组分析已被广泛用于表征细胞状态，但它依赖于第二链互补DNA（CDNA）合成来生成初始材料进行文库制备。在这里，我们使用细菌转座酶TN5，该转座酶TN5越来越多地用于各种高通量DNA分析中，用于构建没有第二链合成的RNA-SEQ库。我们表明，TN5转座体可以随机结合RNA/DNA异链化合物，并在逆转录后直接在RNA上添加测序适配器。该方法是杂音RNA-DNA-HYBRID（雪利酒）的多功能和可扩展性。雪利酒接受从大量RNA到单个细胞的各种起始物质。雪利酒提供了极大的简化协议，并与现行的RNA-seq方法相比，具有更高的可重复性和GC均匀性的结果。
Transcriptome profiling by RNA sequencing (RNA-seq) has been widely used to characterize cellular status, but it relies on second-strand complementary DNA (cDNA) synthesis to generate initial material for library preparation. Here we use bacterial transposase Tn5, which has been increasingly used in various high-throughput DNA analyses, to construct RNA-seq libraries without second-strand synthesis. We show that Tn5 transposome can randomly bind RNA/DNA heteroduplexes and add sequencing adapters onto RNA directly after reverse transcription. This method, Sequencing HEteRo RNA-DNA-hYbrid (SHERRY), is versatile and scalable. SHERRY accepts a wide range of starting materials, from bulk RNA to single cells. SHERRY offers a greatly simplified protocol and produces results with higher reproducibility and GC uniformity compared with prevailing RNA-seq methods.