Protocol for affordable and efficient profiling of nascent RNAs in bread wheat using GRO-seq

Yan Chen; Jiafu Zhu; Yilin Xie; Zijuan Li; Yijing Zhang; Min Liu; Zhicheng Dong

doi:10.1016/j.xpro.2022.101657

Protocol for affordable and efficient profiling of nascent RNAs in bread wheat using GRO-seq

STAR Protoc. 2022 Sep 16;3(3):101657. doi: 10.1016/j.xpro.2022.101657. Epub 2022 Sep 2.

Authors

Yan Chen¹, Jiafu Zhu¹, Yilin Xie², Zijuan Li², Yijing Zhang³, Min Liu¹, Zhicheng Dong¹

Affiliations

¹ Guangdong Provincial Key Laboratory of Plant Adaptation and Molecular Design, Guangzhou Key Laboratory of Crop Gene Editing, Innovative Center of Molecular Genetics and Evolution, School of Life Sciences, Guangzhou University, Guangzhou 510006, China.
² National Key Laboratory of Plant Molecular Genetics, CAS Center for Excellence in Molecular Plant Sciences, Shanghai Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China.
³ State Key Laboratory of Genetic Engineering, Collaborative Innovation Center of Genetics and Development, Department of Biochemistry, Institute of Plant Biology, School of Life Sciences, Fudan University, Shanghai 200438, China.

Abstract

Exorbitant sequencing cost is one of the main obstacles limiting the widespread application of Global Run-On sequencing (GRO-seq) to detect transcriptional activity. Here, we describe a more efficient and affordable protocol for GRO-seq that incorporates an rRNA removal step after nuclear RNA isolation and before nascent RNA immunoprecipitation. We have successfully applied this protocol to profile enhancer transcription in allohexaploid bread wheat and increased the proportion of valid data by 20 times. For complete details on the use and execution of this protocol, please refer to Xie et al. (2022).

Keywords: Bioinformatics; Molecular biology; Plant sciences; RNAseq; Sequence analysis; Sequencing.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Bread*
RNA / genetics
RNA, Nuclear
Sequence Analysis, RNA / methods
Triticum* / genetics

Substances

RNA, Nuclear
RNA