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茶树原位转化方法的建立
引用本文:周承哲,常笑君,朱晨,程春振,陈裕坤,赖钟雄,林玉玲,郭玉琼. 茶树原位转化方法的建立[J]. 生物技术通报, 2022, 0(2): 263-268
作者姓名:周承哲  常笑君  朱晨  程春振  陈裕坤  赖钟雄  林玉玲  郭玉琼
作者单位:福建农林大学园艺学院;福建农林大学茶产业研究院/茶学福建省高校重点实验室;福建农林大学园艺植物生物工程研究所
基金项目:福建农林大学“双一流”建设科技创新能力提升培育计划(KSYLP004);6.18协同创新院茶产业技术分院专项(K1520001A);福建农林大学茶产业链科技创新与服务体系建设项目(K1520005A01);茶树种质资源生化分析与生物学研究专项(K1518023A);福建农林大学乡村振兴茶产业技术服务项目(11899170145)。
摘    要:解决茶树缺乏高效、稳定的遗传转化体系问题,建立了一种基于根癌农杆菌介导的茶树原位转化转基因方法,为茶树基因功能研究和种质创新奠定坚实的基础。以茶树品种‘铁观音’、‘白叶1号’和‘龙井43’的实生幼苗为受体材料,进行去顶芽和腋芽处理。利用根癌农杆菌菌液侵染实生苗伤口,通过抗性筛选获得再生芽,经分子生物学鉴定后,采用短穗扦插法获得茶树转基因植株。结果表明,再生芽中GUS(β-glucuronidase)和HYG(hygromycin)标记基因经多次PCR检测均为阳性,经测序验证PCR产物序列与标记基因序列一致。‘铁观音’、‘白叶1号’和‘龙井43’的转化率分别为8.14%、2.99%和2.53%。建立了不依赖茶树组织培养的原位转化转基因体系,具有操作简便、转化率高、成本低、试验周期短的特点。

关 键 词:茶树  原位转化  根癌农杆菌

Establishment of an Efficient in planta Transformation Method for Camellia sinensis
ZHOU Cheng-zhe,CHANG Xiao-jun,ZHU Chen,CHENG Chun-zhen,CHEN Yu-kun,LAI Zhong-xiong,LIN Yu-ling,GUO Yu-qiong. Establishment of an Efficient in planta Transformation Method for Camellia sinensis[J]. Biotechnology Bulletin, 2022, 0(2): 263-268
Authors:ZHOU Cheng-zhe  CHANG Xiao-jun  ZHU Chen  CHENG Chun-zhen  CHEN Yu-kun  LAI Zhong-xiong  LIN Yu-ling  GUO Yu-qiong
Affiliation:(College of Horticulture,Fujian Agriculture and Forestry University,Fuzhou 350002;Key Laboratory of Tea Science in Universities of Fujian Province,Institute of Tea Industry Research,Fujian Agriculture and Forestry University,Fuzhou 350002;Institute of Horticultural Biotechnology,Fujian Agriculture and Forestry University,Fuzhou 350002)
Abstract:To address the lack of an efficient and stable genetic transformation system for tea trees,an in planta transformation transgenic method based on Agrobacterium tumefaciens-mediated transformation of tea trees was established,laying a solid foundation for gene function research and germplasm innovation of tea tree.The seedlings of tea cultivar ‘Tieguanyin’,‘Baiye1’,and ‘Longjing43’ were used as receptor plants,and the apical bud and axillary bud were removed.Using the A.tumefaciens broth infecting the wounds of the seedling,regenerative buds were obtained through resistance screening.After molecular biological identification,transgenic plants were obtained by short-ear cutting method.The results demonstrated that the GUS(-glucuronidase)and HYG(hyglymycin)labeled genes were tested to be positive with multiple PCR,and the PCR product sequence was consistent with the labeled gene sequence.The transformation efficiency of ‘Tieguanyin’,‘Baiye1’,and ‘Longjing43’ was 8.14%,2.99%,and 2.53%,respectively.This study establishes an in planta transformation method of tea plant that does not rely on tissue culture,and the transformation protocol characterizes of simple operation,high transformation rate,low cost,and short experiment cycle.
Keywords:Camellia sinensis  in planta transformation  agrobacterium tumefaciens
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