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Article|31 Jan 2022|OPEN
Highly efficient activation of endogenous gene in grape using CRISPR/dCas9-based transcriptional activators
Chong Ren1 , Huayang Li1,2 , Yanfei Liu1,2 , Shaohua Li1 , Zhenchang Liang,1 ,
1Beijing Key Laboratory of Grape Sciences and Enology, Key Laboratory of Plant Resource, Institute of Botany, Chinese Academy of Sciences, 20 Nanxincun, Xiangshan, Beijing 100093, China
2University of Chinese Academy of Sciences, 19 Yuquan Rd, Beijing 100049, China
*Corresponding author. E-mail: zl249@ibcas.ac.cn

Horticulture Research 9,
Article number: uhab037 (2022)
doi: https://doi.org/10.1093/hr/uhab037
Views: 180

Received: 05 Jul 2021
Revised: 18 Jan 2022
Accepted: 15 Oct 2021
Published online: 31 Jan 2022

Abstract

Overexpression and knockout (or knockdown) of gene of interest are two commonly used strategies for gene functional study. Clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system-mediated gene knockout has been applied in most plant species, including grapevine. However, CRISPR/dCas9 (deactivated Cas9)-based transcriptional activation is still unreported in fruit crops, although a few studies have been documented in Arabidopsis and rice. Here, we tested two transcriptional activators VP64 and TV for transcriptional activation of endogenous genes in grape. Both the dCas9-VP64 and dCas9-TV systems are efficient enough for transcriptional activation of the UDP-glucose flavonoid glycosyltransferases (UFGT) gene in grape cells. The effectiveness of the dCas9-VP64 system in UFGT activation was about 1.6- to 5.6-fold, while the efficiency of the dCas9-TV system was around 5.7- to 7.2-fold. Moreover, in grapevine plants, highly efficient activation of the cold-responsive transcription factor gene CBF4 was achieved by using the dCas9-TV system. The expression of CBF4 was increased 3.7- to 42.3-fold in transgenic plants. Compared with the wild-type plants, the CBF4-activated plants exhibited lower electrolyte leakage after cold treatment. Our results demonstrate the effectiveness of the dCas9-VP64 and dCas9-TV systems in gene activation in grape, which will facilitate application of transcriptional activation in this economically important species.