摘要
为建立铁皮石斛的SCoT-PCR反应体系,本研究以8个不同增殖代的铁皮石斛组培苗为试验材料,采用正交试验设计,对影响PCR反应体系的2×Taq Master Mix混合液、引物、模板DNA的浓度进行优化,建立了铁皮石斛SCoT-PCR的最佳反应体系,筛选引物,并对1~8代的组培苗进行遗传稳定性鉴定。结果表明:铁皮石斛的SCoT-PCR最佳反应体系(20μL)包含2×Taq Master Mix 10μL、引物0.50μmol/L、模板DNA 40 ng;从60条引物中筛选出19条扩增结果稳定、条带清晰的SCoT引物;所有引物在8个增殖代的16份供试材料中均能扩增出清晰、丰富而稳定的条带,但是所得条带的数量和大小一致,均没有多态性,即16份材料没有发生遗传变异。反应体系的建立可为铁皮石斛种质资源鉴定和遗传多样性研究提供参考,也为其他药用植物组培苗遗传稳定研究提供理论和方法借鉴。
In order to establish the SCoT-PCR system of Dendrobium officinale Kimura et Migo,and explore the genetic stability of tissue culture seedlings of eight different progeny generations,an orthogonal design was used to optimize the concentration of 2×Taq Master Mix,primers,and template DNA that affected the PCR reaction system in this study.The optimal reaction system for SCoT-PCR of Dendrobium officinale Kimura etMigo was estab lished,the primers were screened,and the genetic stability of tissue culture seedlings of 1~8 generations were identified.The results have shown that the optimal SCoT-PCR system(20μL)contains 2×Taq Master Mix 10μL,primer 0.50μmol/L,and template DNA 40 ng.19 SCoT primers which could obtain stable amplification and clear bands were selected from 60.All primers can amplify clear,abundant,and stable bands in 16 test materials of 8 proliferation generations,but the number and size of the bands obtained by all primers are the same and there is no polymorphism.The establishment of this reaction system can provide a possible reference for the identification of germplasm resources and analysis of genetic diversity of Dendrobium officinale Kimura et Migo,as well as a theoretical and methodological reference for the study of genetic stability of tissue culture seedlings of other medicinal plants.
作者
邵征绩
张璐
郑升
陈小军
刘华华
邝曦
龙治坚
韩国辉
Shao Zhengji;Zhang Lu;Zheng Sheng;Chen Xiaojun;Liu Huahua;Kuang Xi;Long Zhijian;Han Guohui(School of Life Science and Engineering,Southwest University of Science and Technology,Mianyang,621010;Fruit Research Institute of Chongqing Academy of Agricultural Sciences,Chongqing,401329;Bamboo Research Institute,Southwest University of Science and Technology,Mianyang,621010)
出处
《分子植物育种》
CAS
北大核心
2021年第5期1606-1613,共8页
Molecular Plant Breeding
基金
国家级大学生创新创业训练计划资助项目(S201910619022,S201910619008X)
四川省大学生创新创业训练计划资助项目(S201910619102,S201910619107)
重庆市农发资金-良种创新重大专项(NKY-2016AA002,NKY-2019AB006)共同资助
