‘云香’水仙ACC合成酶基因NtACS1的克隆及遗传转化  

Molecular Cloning and Its Transformation of ACC Synthase NtACS1 Gene from Narcissus tazetta var.‘Yunxiang'

作　　者：孙申申[1,2] 温秀萍[1,2] 杨菲颖 李梦思[1,2] 李欢[3] 李科[1,2] 陈晓静[1,2] SUN Shenshen;WEN Xiuping;YANG Feiying;LI Mengsi;LI Huan;LI Ke;CHEN Xiaojing(College of Horticulture, Fujian Agriculture and Forestry University, Fuzhou 350002, China;Institute of Genetics and Breeding in Horticultural, Fujian Agriculture and Forestry University, Fuzhou 350002, China;College of The Environment and Ecology, Xiamen University, Xiamen 361102, China)

机构地区：[1]福建农林大学园艺学院,福州350002 [2]福建农林大学园艺植物遗传育种研究所,福州350002 [3]厦门大学环境与生态学院,厦门361102

出　　处：《西北植物学报》2017年第2期250-257,共8页Acta Botanica Boreali-Occidentalia Sinica

基　　金：福建省种业创新与产业化工程项目(K8114001B)

摘　　要：为探究多花水仙ACS基因的序列特征及功能,以‘云香’水仙盛花期花瓣为试验材料,根据‘云香’水仙花朵转录组数据信息,通过RT-PCR方法克隆出1个ACS基因,命名为NtACS1(GenBank KX082936);NtACS1开放阅读框(ORF)长度为552bp,编码183个氨基酸。编码蛋白质分子量约为20.6KDa,理论等电点为6.30,不稳定系数为65.49,属于不稳定的疏水性蛋白。通过qRT-PCR对‘云香’水仙不同时期花瓣和副冠中的NtACS1基因进行了表达分析,得到与‘云香’水仙花朵转录组数据中相同的结果:NtACS1基因在‘云香’水仙花瓣和副冠中的表达都是随着花衰老过程呈现逐渐下降的趋势,且NtACS1基因在花瓣和副冠中的表达峰值都在花苞期,表明NtACS1基因编码的蛋白是在乙烯生物合成途径的系统1发挥催化作用的ACC合成酶。成功构建了NtACS1基因的正义植物表达载体,并通过农杆菌介导法获得8株转基因烟草,PCR和RT-PCR检测显示其中有6株为阳性植株,初步证实NtACS1基因已导入烟草基因组中且在烟草中已表达。该研究结果为进一步分析NtACS1基因的功能和后续转化水仙延长其花期研究奠定了基础。Aimed to study the characteristics and functions of ACS gene,in the present study,we cloned a1-aminocyclopropane-1-carboxylate synthetase gene namedNtACS1(GeneBank KX082936)based on the RNASeq database from the flower ofNarcissus tazettavar.‘Yunxiang’using RT-PCR method.The length of the open reading frame(ORF)of ACS is552bp,encoding183amino acids coupled with a molecular weight of20.6kDa and theoretical isoelectric point of6.30.qRT-PCR analysis showed that the relative expression level of NtACS1both in petals and coronas are decreased gradually along with the aging of flower.Moreover,the expression data ofNtACS1gene were consistent with those obtained by RNA Seq,implied that the NtACS1protein as an ACC synthetase might play a role in the catalytic system1of ethylene biosynthesis.Furthermore,sense plant expression vectors ofNtACS1were successfully constructed with agrobacterium mediated transformation,and6positive transgenic tobacco plants were ultimately obtained.Our current study will lay an experimental foundation for the future application of the genetic transformation to prolong florescence of‘Yunxiang’.

关 键 词：'云香’水仙 基因克隆 表达分析 延长花期 ACS 

分 类 号：Q785[生物学—分子生物学] Q786