黄瓜全雄系A基因的克隆及在杂优育种上的应用

Cucumber (Cucumis sativus L.), which is one of the major vegetable crops, has served as the model system for sex expression in flowering plants. Two main genes responsible for sex determination in cucumber which F/f gene regulates the degree of female flower expression, whereas the M/m gene controls bisexual flower expression have been cloned and both of them encode the 1-aminocyclopropane-1-carboxylic acid syntheses which is key enzyme for ethylene biosynthesis. Using androecious line as the male parent can save the hybrid seed production's costing. However, the genetics and molecular basis of androecious expression in cucumber have rarely been described. A novel androecious gene, which could control the male of cucumber, was discovered by EMS（ethyl methane sulfonate） mutation and designated as a recessive gene by genetic analysis of F2 generation. In this study, we plan to clone the gene through re-sequencing and confirm the function of gene through transgenic technology and co-segregaion of androecious gene SNP(single nucleotide polymorphism) maker with androecious plants in F2 segregating population. Transcriptome sequencing and comparative analysis of cucumber flowers of androecious near-isogenic lines, expression characteristics of the F, M, A gene in the male, female, hermaphrodite flowers via real-time PCR, temporal and spatial characterization of F, M, A gene expression of the male, female, hermaphrodite flowers in situ hybridization are used to analyse the model of F, M and A functions during the development of flowers in cucumber. SNP molecular markers are developed to assist screening new androecious breeding. It will be significant to explain the molecular mechanism of sexual differentiation in cucumber and to facilitate the use of heterosis by manipulating sex expression in cucurbits.

黄瓜（Cucumis sativus L.）是主要蔬菜作物之一，也是研究性别表达的模式植物。黄瓜性别决定主要由基因控制，其中控制单性花的M基因和控制全雌性的F基因已被克隆，均编码乙烯生物合成的关键酶ACC合酶。全雄系可以作为杂种一代的父本，节省制种成本，但目前控制全雄系的基因尚无报道。前期研究中，通过EMS诱变发现了一份全雄系，遗传分析表明全雄系受单隐性基因控制。本研究拟通过重测序克隆得到黄瓜全雄系基因A，并开发SNP标记及通过转基因技术对其功能进行验证；进一步采用转录组测序、Real-time PCR、原位杂交等技术探讨性别决定基因F、M、A互作对黄瓜性型的影响，从而完善黄瓜性别表达调控模型；建立全雄系分子标记辅助选育体系并运用其选育新父本材料。该研究对黄瓜性别决定分子机理的解析和瓜类杂优育种新技术开发具有重要意义

黄瓜（Cucumis sativus L.）是葫芦科的重要经济作物，长期以来一直是性别决定研究的重要模式系统。本项目从EMS（甲基磺酸乙酯）诱变的黄瓜全雄突变材料中克隆得到全雄系基因A（后命名为CsACO2），该基因编码乙烯合成途径中的关键酶ACC氧化酶（CsACO2）。研究发现功能保守区的异义突变P254S能导致CsACO2酶活完全丧失。全雄突变植株茎端的乙烯释放量和ACO总的酶活性也降低。利用原位杂交实验检测CsACO2和CsWIP1的表达模式，结果表明CsACO2在花发育的第2期至第4期心皮处特异表达，与CsACS11在雌花中的表达区域存在重叠，而与CsWIP1的表达区域不重叠，CsACO2通过配合CsACS11控制雌花心皮处乙烯特异性升高来影响花芽持续向雌花方向发育。进一步通过烟草瞬时转化体系、酵母单杂交、凝胶阻滞和黄瓜原生质体的ChIP实验，证明了CsWIP1是调控CsACO2表达的一个重要因子，CsWIP1可以结合CsACO2的启动子从而抑制该基因的表达。CsACO2调控黄瓜性别决定机理的发现，完善了黄瓜性别表达调控模型。此外，本项目还利用Ⅲ-12-8全雄株（ACO2）基因开发分子标记，建立分子标记辅助选择育种体系，并初步探索黄瓜新的制种模式，为黄瓜规模化生产以及瓜类杂优育种新技术开发做出贡献。