苜蓿次生胚遗传转化系统优化  

Optimization of Genetic Transformation of Secondary Somatic Embryo in Alfalfa

作　　者：金太成[1] 王悦 吴艳菊 孟大伟[1] 杨丽萍[1] Jin Taicheng;Wang Yue;Wu Yanju;Meng Dawei;Yang Liping(Key Laboratory for Plant Resources Science and Green Production,The School of Life Science,Jilin Normal University,Siping,136000)

机构地区：[1]吉林师范大学生命科学学院吉林省植物资源科学与绿色生产重点实验室

出　　处：《分子植物育种》2019年第20期6685-6690,共6页Molecular Plant Breeding

基　　金：吉林省教育厅“十三五”科研规划项目(JJKH20180794KJ)资助

摘　　要：稳定高效的遗传转化系统是对苜蓿进行基因改良的重要前提条件。本实验以吉林省适产苜蓿品种为实验材料,对农杆菌介导的次生胚遗传转化系统中的各种关键影响因素进行了优化:胚性愈伤诱导培养基中2,4-D和KT的最适浓度分别为2 mg/L和0.5 mg/L;体胚分化培养基中6-BA和NAA的最适浓度均为0.5 mg/L;该法最适受体基因型为公农系列苜蓿品种;遗传转化过程中,胚性愈伤诱导培养基及体胚分化培养基中添加的卡那霉素浓度分别为75 mg/L和50 mg/L。通过GUS基因表达检测和定量RT-PCR分析表明,外源基因已经顺利整合进苜蓿基因组染色体中,遗传转化效率为20%左右。本实验为培育吉林省转基因苜蓿新品种工作提供了科学依据。Stable transform system is the premise condition for alfalfa genetic improvement. We took several alfalfa varieties suitable planted in Jilin Province as experimental materials. Through infection of alfalfa cotyledon-stage somatic embryos by Agrobacterium strain GV3101, we successfully transformed foreign gene into the genome of alfalfa. We optimized the key factors in this transform system and found that the optimal concentration of 2,4-D and KT in embryonic callus medium is respectively 2 mg/L and 0.5 mg/L;the optimal concentration of6-BA and NAA in somatic embryogenesis medium is both 0.5 mg/L;the optimal genotype of alfalfa in this system is Gongnong series;the optimal concentration of Kanamycin in embryonic callus medium and somatic embryogenesis medium is respectively 75 mg/L and 50 mg/L;by the detection of GUS expression and semi-quantitative PCR analysis of GUS gene, the reporter gene has been successfully transformed in the genome of alfalfa. The whole transformation efficiency is reached into 20%. This experiment provided scientific evidences for developing a new transgenic alfalfa variety suitable for Jilin Province.

关 键 词：苜蓿(Medicago SATIVA L.) 遗传转化 次生胚 

分 类 号：S54[农业科学—作物学]