Salt and osmotic stress can improve the editing efficiency of CRISPR/Cas9-mediated genome editing system in potato

PeerJ

article

Mingwang Ye¹  Mengfan Yao¹  Canhui Li³  Ming Gong¹ 

[1] School of Life Sciences, Yunnan Normal University;Engineering Research Center of Sustainable Development and Utilization of Biomass Energy, Ministry of Education, Yunnan Normal University;Yunnan Key Laboratory of Potato Biology, Yunnan Normal University;Joint Academy of Potato Science, Yunnan Normal University;Key Laboratory of Biomass Energy and Environmental Biotechnology of Yunnan Province, Yunnan Normal University

关键词: Genome editing efficiency;    CRISPR/Cas9;    Salt and osmotic stress;    Potato;    A. rhizogenes;   

DOI  :  10.7717/peerj.15771

学科分类：社会科学、人文和艺术（综合）

来源: Inra

【 摘 要 】

CRISPR/Cas9-mediated genome editing technology has been widely used for the study of gene function in crops, but the differences between species have led to widely varying genome editing efficiencies. The present study utilized a potato hairy root genetic transformation system and incorporated a rapid assay with GFP as a screening marker. The results clearly demonstrated that salt and osmotic stress induced by NaCl (10 to 50 mM) and mannitol (50 to 200 mM) treatments significantly increased the positive rates of genetic transformation mediated by A. rhizogenes and the editing efficiency of the CRISPR/Cas9-mediated genome editing system in potato. However, it was observed that the regeneration of potato roots was partially inhibited as a result. The analysis of CRISPR/Cas9-mediated mutation types revealed that chimeras accounted for the largest proportion, ranging from 62.50% to 100%. Moreover, the application of salt and osmotic stress resulted in an increased probability of null mutations in potato. Notably, the highest rate of null mutations, reaching 37.5%, was observed at a NaCl concentration of 10 mM. Three potential off-target sites were sequenced and no off-targeting was found. In conclusion, the application of appropriate salt and osmotic stress significantly improved the editing efficiency of the CRISPR/Cas9-mediated genome editing system in potato, with no observed off-target effects. However, there was a trade-off as the regeneration of potato roots was partially inhibited. Overall, these findings present a new and convenient approach to enhance the genome editing efficiency of the CRISPR/Cas9-mediated gene editing system in potato.

【 授权许可】

CC BY