乙烯调控罗汉果性别的细胞学及分子遗传研究

Siraitia grosvenorii is a kind of precious medical and edible economic crop in China, characterized by male and female reproductive organs occurring on different individuals. Therefore, artificial pollination should be conducted to get enough fruits, which wasted plenty of time, labor and cost,and caused products supply falling short of market demand. That discovering the genetic base of sex would be the key to solving this problem from fountainhead of reeding. Several key research aspects would be carried out as follows:bisexual flower was induced by ethylene inhibitor; microstructure between male and female bud/flower of siraitia grosvenorii would be observed by comparison from floral initiation to the formation of siraitia grosvenorii; for male and female reproductive organs at key development stages, studies of transcriptome and digital gene expression profile (DGE) would be analyzed, followed by their function analysis of key genes were performed to reveal the molecular genetics mechanism of sex determination and expression; then developmental biology, quality and other important characters was nvestigated for transgenic generation of sex modified siraitia grosvenorii.

罗汉果是我国重要特色药食兼用经济作物，因雌雄异株需要大量人工辅助授粉，生产成本高，产品供不应求，是其产业化发展中存在的主要问题，明确性别形成的遗传基础是从育种源头解决问题的关键。拟喷施乙烯抑制剂，诱导罗汉果形成两性花植株；比较研究从花芽分化到配子体形成过程中与自然状态雌雄异株罗汉果花/芽发育的细胞学差异；研究性别分化和性器官形成关键时期转录组和表达谱的差异，研究与乙烯调控相关的关键基因的功能，明确与乙烯调控相关的性别决定和表达的分子遗传；研究罗汉果性别修饰后代的发育生物学、品质等主要性状表现等。

罗汉果[Siraitia grosvenorii]为广西特色药食两用植物，因雌雄异株栽培中需要人工授粉，生产成本高。因此，研究调控罗汉果的花芽发育过程和花器官形成关键基因，不仅对揭示其性器官的形态建成具有重要理论意义，而且对应用于遗传育种以实现性别逆转而免除人工授粉具重要实践意义。项目利用AgNO3 诱导罗汉果雌株形成两性花，对其进行花器官形态学、细胞学、组织化学研究；分析雌、雄花及诱导两性花在转录组水平基因差异表达；克隆WIP、ACS1、ACS3、 ACS10 和 ACS12 5个基因cDNA和DNA全长，分析基因生物信息学及在罗汉果雄株、雌株以及诱导两性花在不同时期不同器官相对表达量；检测银离子处理前后雌株花芽的甲基化状态并进行MSAP 分析。 结果如下：.（1）80~400mg/L AgNO3均能诱导形成两性花，但诱导形成的两性花花药无花粉粒形成；各浓度AgNO3诱导的雄蕊缺乏淀粉粒和蛋白质积累，花粉母细胞时期退化。AgNO3影响子房、胚珠、胚囊母细胞形态。.（2）通过两个时期雌花、雄花和诱导两性花的转录组差异分析总结为如下：①罗汉果性别分化表现偏向性基因表达规律；②罗汉果性别分化受乙烯激素信号等环境信息加工调控；③转录因子和可变剪切在罗汉果性别分化中发挥重要调节作用；④罗汉果性别分化还与碳水化合物、苯丙素代谢、氨基酸代谢等多个代谢密切相关。.（3）获得 SgWIP、SgACS1、SgACS3、SgACS10 和 SgACS12 基因cDNA和DNA全长；发现SgWIP1 、SgACS1 、SgACS 10 在生殖部位的表达量高于营养器官，SgWIP1 SgACS1和 SgACS 10表达与外源AgNO3 正相关；SgACS3受 AgNO3 负调控表达。.（4）诱导两性花甲基化水平高于雌花；两性花中新增的甲基化位点多为全甲基化， AgNO3 可能通过诱导雌花 DNA 无甲基化位点发生全甲基化，影响基因表达。