番茄遗传转化与筛选鉴定的改进研究  

Optimization of Tomato Genetic Transformation and Identification

作　　者：祁春芳 王婷 刘霞宇 石晓雯 于治芹 刘世芳 梁建萍[2] 贾小云[2] QI Chunfang;WANG Ting;LIU Xiayu;SHI Xiaowen;YU Zhiqn;LIU Shifang;LIANG Jianping;JIA Xiaoyun(Sangganhe Poplar Forest Experiment Bureau,Datong Shanxi 037006,China;College of Life Seience,Shanxi Agricultural University,Taigu Shanxi 030801 ,China)

机构地区：[1]山西省桑干河杨树丰产林实验局,山西大同037006 [2]山西农业大学生命科学学院,山西太谷030801

出　　处：《西北农业学报》2018年第4期554-561,共8页Acta Agriculturae Boreali-occidentalis Sinica

基　　金：国家自然科学基金(31101555);山西农业大学研究生创新项目(20143058);山西农业大学科技创新基金(2018);山西省高等学校优秀青年学术带头人项目(2012);山西人才引进与开发专项(614191);山西省农业科技成果转化和推广示范项目(2018-5)~~

摘　　要：将At-pri-miR828基因的过表达载体pC2300-pOT2-At-pri-miR828通过农杆菌侵染法转入番茄(Solanum lycopersicum L.)品种‘Ailsa Craig’为例,来对番茄的遗传转化过程中番茄种子消毒处理、无菌苗培养、转基因阳性植株的Kan筛选浓度、PCR检测等方法进行研究,旨在探索一套快速高效的番茄遗传转化及鉴定方法。研究发现:采用C_2H_5OH、Na_3PO_3及NaClO分别浸泡及无菌水反复冲洗的种子消毒方法,可使污染率减少到5%以下;种子震荡培养法可以提高发芽率并促进种苗的生长势一致;筛选转基因阳性植株的Kan最佳质量浓度为100mg·L^(-1);100mg·L^(-1)的Kan溶液浇灌蛭石和在MS培养基中添加Kan的方法都可以用来筛选转基因番茄。最终Kan筛选和PCR检测等鉴定方法显示番茄的遗传转化效率为32.5%。To explore a rapid and efficient method for tomato genetic transformation and identification,this research studied conditions of seed disinfection,seedling culture,kanamycin screening of transgenic plants,PCR conformation,etc.,taken the overexpression vector pC2300-pOT22-At-primiR828 transferred into the cotyledon explants of tomato（Solanum lycopersicum L.‘Ailsa Craig＇）as an example.The results showed that the contamination rate of tomato seeds was decreased to below 5% when they were disinfected with C2H5OH,Na3PO3 and NaClO,respectively followed by sterile water washing more times.The seed-shaking culture method can improve the germination rate and uniformity degree of seedlings.The Kan concentration of 100 mg·L^-1 was optimal for screening transgenic tomatoes.The transgenic tomatoes can be screened by directly adding 100 mg·L^-1 Kan into vermiculite or MS medium.The genetic transformation efficiency of tomato screened by Kan resistance and PCR detection was 32.5%.

关 键 词：番茄 种子消毒 遗传转化 Kan筛选 

分 类 号：S641.2[农业科学—蔬菜学]