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Article|17 Nov 2023|OPEN
Development of virus-induced genome editing methods in Solanaceous crops
Seo-Young Lee1,3 , Bomi Kang2,3 , Jelli Venkatesh1 , Joung-Ho Lee1 , Seyoung Lee1 , Jung-Min Kim2 and Seungki Back1 , Jin-Kyung Kwon1 , Byoung-Cheorl Kang,1,2 ,
1Department of Agriculture, Forestry and Bioresources, Research Institute of Agriculture and Life Sciences, Plant Genomics Breeding Institute, College of Agriculture and Life Sciences, Seoul National University, Seoul 08826, Republic of Korea
2Interdisciplinary Program in Agricultural Biotechnology, College of Agriculture and Life Sciences, Seoul National University, Seoul 08826, Republic of Korea
3These authors contributed equally to this manuscript
*Corresponding author. E-mail: bk54@snu.ac.kr

Horticulture Research 11,
Article number: uhad233 (2024)
doi: https://doi.org/10.1093/hr/uhad233
Views: 27

Received: 08 Jun 2023
Accepted: 01 Nov 2023
Published online: 17 Nov 2023

Abstract

Genome editing (GE) using CRISPR/Cas systems has revolutionized plant mutagenesis. However, conventional transgene-mediated GE methods have limitations due to the time-consuming generation of stable transgenic lines expressing the Cas9/single guide RNA (sgRNA) module through tissue cultures. Virus-induced genome editing (VIGE) systems have been successfully employed in model plants, such as Arabidopsis thaliana and Nicotiana spp. In this study, we developed two VIGE methods for Solanaceous plants. First, we used the tobacco rattle virus (TRV) vector to deliver sgRNAs into a transgenic tomato (Solanum lycopersicum) line of cultivar Micro-Tom expressing Cas9. Second, we devised a transgene-free GE method based on a potato virus X (PVX) vector to deliver Cas9 and sgRNAs. We designed and cloned sgRNAs targeting Phytoene desaturase in the VIGE vectors and determined optimal conditions for VIGE. We evaluated VIGE efficiency through deep sequencing of the target gene after viral vector inoculation, detecting 40.3% and 36.5% mutation rates for TRV- and PVX-mediated GE, respectively. To improve editing efficiency, we applied a 37°C heat treatment, which increased the editing efficiency by 33% to 46% and 56% to 76% for TRV- and PVX-mediated VIGE, respectively. To obtain edited plants, we subjected inoculated cotyledons to tissue culture, yielding successful editing events. We also demonstrated that PVX-mediated GE can be applied to other Solanaceous crops, such as potato (Solanum tuberosum) and eggplant (Solanum melongena). These simple and highly efficient VIGE methods have great potential for generating genome-edited plants in Solanaceous crops.