葡萄ACO家族基因过量表达对番茄乙烯释放速率的影响  

Over-expression of 1-aminocyclopropane-1-carboxylic acid oxidase Genes(ACO) from Vitis vinifera in Tomato(Solanum locopersicum) and Its Effects on Ethylene Release Rates

作　　者：蔡涛[1] 叶霞[2] 马媛春 王小龙[1] 陈飞[1] 程宗明[1] 

机构地区：[1]南京农业大学园艺学院,南京210095 [2]河南农业大学园艺学院,郑州450002

出　　处：《农业生物技术学报》2013年第9期1037-1044,共8页Journal of Agricultural Biotechnology

基　　金：国家自然科学基金项目(No.31201605)

摘　　要：乙烯是启动和促进果蔬成熟和衰老的内源激素,ACC氧化酶(1-aminocyclopropane-1-carboxylic acid oxidase,ACO)是乙烯生物合成中的一个关键限速酶。葡萄ACO基因拥有一个基因家族,其中VvACO1和VvACO2基因在果实转色时期出现转录高峰,是葡萄果实发育过程中造成乙烯含量发生变化的关键基因。为验证葡萄(Vitis vinifera)ACO家族基因过量表达对番茄(Solanum locopersicum)乙烯释放速率的影响,本研究通过根癌农杆菌(Agrobacterium tumefaciens)介导法将葡萄ACO基因导入番茄基因组中,进行了PCR和RT-PCR分子检测,并对分子检测阳性的转基因番茄植株叶片的ACO酶活性和乙烯释放速率进行了分析。RT-PCR检测表明,在3个VvACO1、4个VvACO2和6个VvACO3转基因番茄植株中目的基因均有一定的表达。所有转基因番茄植株的ACO酶活性和乙烯释放速率均比对照有所提高,其中以VvACO1基因的过量表达对叶片ACO酶活和乙烯释放速率增加的幅度最大,VvACO3基因的过量表达对果实的ACO酶活和乙烯释放速率增加的幅度最大,而转化VvACO2基因的番茄植株ACO酶活性和乙烯释放速率最低,比对照略有升高,且植株呈现矮化现象。本研究通过观察转基因番茄的生长状况以及测定转基因番茄的生理指标,探讨葡萄ACO家族基因的过量表达对番茄的影响,来预测葡萄ACO基因的初步功能。Ethylene is an important plant endogenous hormone which controls fruit ripening and senescence, and 1-aminocyclopropane-1-carboxylic acid oxidase（ACO） is one of the key rate-determining enzymes in the biosynthesis of ethylene. VvACOs is coded by a small gene family, and the highest expression of VvAC01 and VvAC02, which are key genes regulating ethylene production changes during berry fruit development, is observed in the stage of veraison. In order to verify the influences of over-expression of VvACOs on ethylene release rates in tomato（Solanum locopersicum）, Grapevine 1-aminocyclopropane-1-carboxylic acid oxidase （VvACO） genes were introduced into tomato genome by Agrobacturiura mediated transformation method. The transgenic tomato plants were further verified with PCR and RT-PCR analysis, and ACO enzyme activities and ethylene release rates in the leaves of transgenic tomato plants were measured by gas chromatography. RT- PCR analysis indicated that three, four and six transgenic tomato lines with over-expressing VvACO1, VvAC02 and VvAC03 were obtained, respectively. The ACO enzyme activities and ethylene release rates in the transgenic plants were higher than those in non-transgenic tomato leaves and fruits. The highest ACO enzyme activities and ethylene release rates in the leaves and fruits of tomato transgenic lines with over-expressing VvACO1 and VvAC03 were observed, respectively. The transgenic tomato plants which over-expressing both VvACO1 and VvAC03 were normal morphological growth. The lowest ACO enzyme activities and ethylene release rates were observed in the tomato transgenic lines with over-expressing VvAC02, and the VvAC02 over- expressing plants were shown dwarf phenotype. Transgenic tomatoes that over-expressed of the each of the VvACO genes were analyzed for their growth physiological indexs, ethylene release rates, and ACO enzyme activies. Based on these results, functions of VvACO ~enes were preliminarily predicted.

关 键 词：葡萄 ACO基因 乙烯释放速率 过量表达 农杆菌介导遗传转化 

分 类 号：S663.1[农业科学—果树学]