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Short Communication | Open Access

Systematic identification of endogenous RNA polymerase Ⅲ promoters for efficient RNA guide-based genome editing technologies in maize

Xiantao Qia,b,1, Le Donga,1, Changlin Liua, Long Maoa, Fang Liua, Xin Zhangb, Beijiu Chengb, Chuanxiao Xiea( )
Institute of Crop Science, Chinese Academy of Agricultural Sciences, Beijing 100081, China
Anhui Agricultural University, Anhui Province, Hefei 230036, Anhui, China

1 These authors contributed equally to this work.

Peer review under responsibility of Crop Science Society of China and Institute of Crop Science, CAAS.

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Abstract

Single-guide RNA (sgRNA) is one of the two core components of the CRISPR (clustered regularly interspaced short palindromic repeat)/Cas (CRISPR-associated) genome-editing technology. We established an in vitro Traffic Light Reporter (TLR) system, which is designated as the same colors as traffic lights such as green, red and yellow were produced in cells. The TLR can be readily used in maize mesophyll protoplast for a quick test of promoter activity. The TLR assay indicates the variation in transcription activities of the seven Pol Ⅲ promoters, from 3.4% (U6-1) to over 21.0% (U6-6). The U6-2 promoter, which was constructed to drive sgRNA expression targeting the ZmWx1 gene, yielded mutation efficiencies ranging from 48.5% to 97.1%. Based on the reported and unpublished data, the in vitro TLR assay results were confirmed to be a readily system and may be extended to other plant species amenable to efficient genome editing via CRISPR/Cas. Our efforts provide an efficient method of identifying native Pol Ⅲ-recognized promoters for RNA guide-based genome-editing systems in maize.

Keywords

Genome editingCRISPR/CasMaizeRNA polymerase Ⅲ promoters

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The Crop Journal
Volume 6 Issue 3,
June 2018
Pages 314-320
DOI: 10.1016/j.cj.2018.02.005
Cite this article:
Qi X, Dong L, Liu C, et al. Systematic identification of endogenous RNA polymerase Ⅲ promoters for efficient RNA guide-based genome editing technologies in maize. The Crop Journal, 2018, 6(3): 314-320. https://doi.org/10.1016/j.cj.2018.02.005

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Received: 27 December 2017
Revised: 31 January 2018
Accepted: 01 March 2018
Published: 03 April 2018
© 2018 Crop Science Society of China and Institute of Crop Science, CAAS.

(http://creativecommons.org/licenses/by-nc-nd/4.0/).
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