农杆菌介导薄皮甜瓜遗传转化体系建立

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英文篇名：Establishment of Agrobacterium mediated genetic transformation system of muskmelon 作者：栾非时 ; 白晶 ; 朱子成 ; 高鹏 ; 刘识 ; 吴川 ; 樊超 ; 吕双雪 英文作者：LUAN Feishi;BAI Jing;ZHU Zicheng;GAO Peng;LIU Shi;WU Chuan;FAN Chao;LV Shuangxue;Key Laboratory of Biology and Genetic Improvement Horticultural Crops (Northeast Region), Ministry of Agriculture;School of Horticulture and Landscape Architecture, Northeast Agricultural University; 关键词：薄皮甜瓜 ; 子叶节 ; 再生 ; 遗传转化 英文关键词：muskmelon;;cotyledonary node;;regeneration;;genetic transformation 中文刊名：东北农业大学学报 英文刊名：Journal of Northeast Agricultural University 机构：农业部东北地区园艺作物生物学与种质创制重点实验室;东北农业大学园艺园林学院; 出版日期：2019-01-15 10:37 出版单位：东北农业大学学报 年：2019 期：01 基金：国家西甜瓜产业技术体系项目(CARS-25);; 国家自然科学基金项目(31772331) 语种：中文; 页：14-21 页数：8 CN：23-1391/S ISSN：1005-9369 分类号：S652

摘要

以薄皮甜瓜"M15"和"M43"子叶节为外植体,研究激素浓度、卡那霉素浓度和消毒时间等对甜瓜子叶节再生及农杆菌介导遗传转化的影响。结果表明,诱导甜瓜子叶节不定芽分化最佳激素组合为0.5 mg·L~(-1)6-BA+0.05 mg·L~(-1)IAA,芽伸长激素为0.05 mg·L~(-1)6-BA,生根激素为0.1 mg·L~(-1)IAA。农杆菌介导甜瓜转化最佳条件为预培养48 h,OD_(600)=0.6农杆菌菌液侵染15 min,黑暗条件下共培养48～72 h,头孢抑菌浓度500 mg·L~(-1),卡那霉素浓度75 mg·L~(-1)。PCR检测卡那霉素抗性苗,验证外源基因已整合进入甜瓜基因组,"M15"和"M43"抗性植株阳性率分别为84.2%和77.4%。
        Using cotyledon nodes of muskmelon "M15" and "M43" as explants, the factors that affected melon cotyledon regeneration and the efficiency of genetic transformation were studied, such as hormone concentration, kanamycin concentration, and disinfect time. The results showed that the best hormone combination of adventitious bud differentiation in cotyledons of muskmelon was 0.5 mg· L~(-1)6-BA+0.05 mg· L~(-1) IAA, shoot elongation hormone was 0.05 mg· L~(-1)6-BA and rooting hormone was 0.1 mg· L~(-1) IAA. The best condition for Agrobacterium mediated transformation of muskmelon was pre-culture for48 h, OD_(600)=0.6 Agrobacterium microbial infection for 15 min, co-cultivate for 48-72 h in the dark,cephalosporins bacteriostatic concentration of 500 mg· L~(-1), kanamycin concentration of 75 mg· L~(-1). It was originally proved by PCR that expression cassettes had been transferred into the genome, the positive rate of "M15" and "M43" were 84.2% and 77.4%, respectively.

引文

[1]王喜庆.黑龙江省西瓜甜瓜生产现状、存在的问题和对策[J].中国瓜菜,2008(2):53-54.
    [2]Fang G,Grumet R.Agrobacterium tumefaciens mediated transformation and regeneration of muskmelon plants[J].Plant Cell Rep,1990(9):160-164.
    [3]Choi J Y,Shin J S,Chung Y S,et al.An efficient selection and regeneration protocol for Agrobacterium-mediated transformation of oriental melon(Cucumis melo L.var.makuwa)[J].Plant Cell Tiss Organ Cult,2012,110:133-140.
    [4]郝金凤,荆培培,张丽,等.应用农杆菌介导的生长点转化方法建立甜瓜遗传转化技术[J].华北农学报,2014,29(2):116-120.
    [5]李蕾,孟永娇,张璐.黄瓜遗传转化体系优化的研究[J].华北农学报,2015,30(5):115-121.
    [6]Lin C Y,Ku H M,Chiang Y H,et al.Development of transgenic watermelon resistant to Cucumber mosaic virus and watermelon mosaic virus by using a single chimeric transgene construct[J].Transgenic Res,2012,21:983-993.
    [7]王慧中,赵培洁,周晓云.转基因甜瓜植株的获得及其抗病性[J].植物保护学报,2000,27(2):126-130.
    [8]樊继德,何启伟,王秀峰,等.甜瓜反义酸性转化酶基因对甜瓜的遗传转化[J].园艺学报,2007,34(3):677-682.
    [9]Nu?ez-Palenius H G,Cantliffe D J,Huber D J,et al.Transformation of a muskmelon"Galia"hybrid parental line(Cucumis melo L.var.reticulatus Ser.)with an antisense ACC oxidase gene[J].Plant Cell Rep,2006,25:198-205.
    [10]付秋实,谭明明,王婢,等.不同甜瓜品种再生体系的比较研究[J].中国瓜菜,2015,28(2):5-8.
    [11]肖守华,赵善仓,王崇启,等.厚皮甜瓜高效再生体系的建立[J].山东农业科学,2007(4):35-39.
    [12]付秋实,曹芸运,谭明明,等.薄皮甜瓜再生体系的优化[J].中国瓜菜,2014,27(2):16-19.
    [13]武云鹏,张若纬,彭冬秀,等.薄皮甜瓜‘花雷’再生体系的建立[J].中国瓜菜,2018,31(11):36-39.
    [14]黄永红,陆璐,陶兴林,等.甜瓜品种GT-1植株再生体系的建立[J].果树学报,2006,23(5):740-744.
    [15]张慧君,高鹏,栾非时.甜瓜自交系离体子叶节再生体系的建立[J]江苏农业科学,2012,40(3):50-52.
    [16]颜雪.根癌农杆菌介导R-Fom-2基因转化新疆甜瓜的研究[D].乌鲁木齐:新疆大学,2008.
    [17]Yadav R C,Salah M T,Grumet R.High frequency shoot regeneration from leaf explants of muskmelon[J].Plant Cell Tiss Organ Cult,1996,45:207-214.
    [18]杨兰,张宝燕.不同的抗生素对农杆菌EHA105(codA)和农杆菌EHA105(betA)的抑制作用[J].山东林业科技,2011(2):9-13.
    [19]高宁宁,张显,郭蔼光,等.抗菌肽基因转化厚皮甜瓜的研究[J].西北植物学报,2011,31(11):2158-2164.