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利用CRISPR/Cas13a编辑技术获得马铃薯抗PVX植株
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引用本文:涂振,钟子旸,陈汝豪,杨曼华,詹晓慧,陈家茹,马恢,聂碧华.利用CRISPR/Cas13a编辑技术获得马铃薯抗PVX植株.植物保护学报,2024,51(1):211-218
DOI:10.13802/j.cnki.zwbhxb.2024.2022112
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作者单位E-mail
涂振 华中农业大学园艺林学学院, 果蔬园艺作物种质创新与利用全国重点实验室, 农业农村部马铃薯生物学与生物技术重点实验室, 武汉 430070  
钟子旸 华中农业大学园艺林学学院, 果蔬园艺作物种质创新与利用全国重点实验室, 农业农村部马铃薯生物学与生物技术重点实验室, 武汉 430070
恒敬合创生物医药 (浙江) 有限公司, 浙江 嘉兴 314000 
 
陈汝豪 华中农业大学园艺林学学院, 果蔬园艺作物种质创新与利用全国重点实验室, 农业农村部马铃薯生物学与生物技术重点实验室, 武汉 430070  
杨曼华 华中农业大学园艺林学学院, 果蔬园艺作物种质创新与利用全国重点实验室, 农业农村部马铃薯生物学与生物技术重点实验室, 武汉 430070  
詹晓慧 湖北大学生命科学学院, 省部共建生物催化与酶工程国家重点实验室, 武汉 430062  
陈家茹 华中农业大学园艺林学学院, 果蔬园艺作物种质创新与利用全国重点实验室, 农业农村部马铃薯生物学与生物技术重点实验室, 武汉 430070  
马恢 张家口市农业科学院马铃薯研究所, 河北 张家口 075000  
聂碧华 华中农业大学园艺林学学院, 果蔬园艺作物种质创新与利用全国重点实验室, 农业农村部马铃薯生物学与生物技术重点实验室, 武汉 430070 nbihua@mail.hzau.edu.cn 
中文摘要:为探究利用CRISPR/Cas13a系统获得抗马铃薯X病毒(potato virus X,PVX)马铃薯的可行性,通过设计靶向PVX中TGBp1基因的小向导RNA(small guide RNA,sgRNA),构建CRISPR/Cas13a基因编辑载体,以马铃薯栽培种Désirée为受体材料进行稳定遗传转化,并通过机械摩擦接种法鉴定转基因植株对PVX的抗性。结果显示,成功构建了靶向PVX的PVX-Cas13a载体,并获得了表达Cas13a的转基因马铃薯植株,对其中3个高表达Cas13a的株系进行PVX抗性鉴定,发现在接种PVX 10~20 d后,转基因植株系统叶上无明显发病症状,且PVX积累量均显著低于未接种PVX对照。表明靶向PVX的CRISPR/Cas13a系统能够有效抑制该病毒的积累,这为马铃薯抗PVX育种提供了一条有效策略。
中文关键词:马铃薯  基因编辑  CRISPR/Cas13a  马铃薯X病毒  抗性鉴定
 
Generation of PVX resistant potato plants by CRISPR/Cas13a editing technology
Author NameAffiliationE-mail
Tu Zhen Key Laboratory of Potato Biology and Biotechnology, Ministry of Agriculture and Rural Affairs
National Key Laboratory for Germplasm Innovation & Utilization of Horticultural Crops
College of Horticulture & Forestry Sciences, Huazhong Agricultural University, Wuhan 430070, Hubei Province, China 
 
Zhong Ziyang Key Laboratory of Potato Biology and Biotechnology, Ministry of Agriculture and Rural Affairs
National Key Laboratory for Germplasm Innovation & Utilization of Horticultural Crops
College of Horticulture & Forestry Sciences, Huazhong Agricultural University, Wuhan 430070, Hubei Province, China
Virtu Pharmako (Zhejiang) Company Limited, Jiaxing 314000, Zhejiang Province, China 
 
Chen Ruhao Key Laboratory of Potato Biology and Biotechnology, Ministry of Agriculture and Rural Affairs
National Key Laboratory for Germplasm Innovation & Utilization of Horticultural Crops
College of Horticulture & Forestry Sciences, Huazhong Agricultural University, Wuhan 430070, Hubei Province, China 
 
Yang Manhua Key Laboratory of Potato Biology and Biotechnology, Ministry of Agriculture and Rural Affairs
National Key Laboratory for Germplasm Innovation & Utilization of Horticultural Crops
College of Horticulture & Forestry Sciences, Huazhong Agricultural University, Wuhan 430070, Hubei Province, China 
 
Zhan Xiaohui State Key Laboratory of Biocatalysis and Enzyme Engineering, School of Life Sciences, Hubei University, Wuhan 430062, Hubei Province, China  
Chen Jiaru Key Laboratory of Potato Biology and Biotechnology, Ministry of Agriculture and Rural Affairs
National Key Laboratory for Germplasm Innovation & Utilization of Horticultural Crops
College of Horticulture & Forestry Sciences, Huazhong Agricultural University, Wuhan 430070, Hubei Province, China 
 
Ma Hui Institute of Potato Research, Zhangjiakou Academy of Agricultural Sciences, Zhangjiakou 075000, Hebei Province, China  
Nie Bihua Key Laboratory of Potato Biology and Biotechnology, Ministry of Agriculture and Rural Affairs
National Key Laboratory for Germplasm Innovation & Utilization of Horticultural Crops
College of Horticulture & Forestry Sciences, Huazhong Agricultural University, Wuhan 430070, Hubei Province, China 
nbihua@mail.hzau.edu.cn 
Abstract:To investigate the feasibility of utilizing the CRISPR/Cas13a system to confer resistance against potato virus X (PVX) in Solanum tuberosum, a targeted small guide RNA (sgRNA) was designed to disrupt the TGBp1 gene of PVX and was employed in constructing a CRISPR/Cas13a geneediting vector. The potato cultivar Désirée was chosen for stable genetic transformation. The results showed that transgenic potato lines expressing Cas13a were successfully developed and subsequently subjected to assessment for PVX resistance via mechanical inoculation. Among the transgenic lines, three demonstrated robust Cas13a expression and exhibited resistance to PVX. These resistant lines displayed an absence of disease symptoms on systemically infected leaves and showcased a substantial reduction in PVX accumulation between 10 to 20 days post-inoculation. These findings underscore the potency of the CRISPR/Cas13a system in effectively targeting PVX, thereby impeding virus accumulation. This study delineated the promising potential of harnessing CRISPR/Cas13a as a viable strategy for breeding PVX-resistant potatoes.
keywords:potato  gene editing  CRISPR/Cas13a  potato virus X  resistance identification
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