芝麻细胞色素P450超家族的成员鉴定及SiCYP703A2的功能验证  

Identification of members of cytochrome P450 superfamily and functional characterization of SiCYP703A2 gene in sesame

作　　者：李田雨 杨远霄 刘红艳[1] 汪魏 周芳[1] 周婷[1] 赵应忠[1] LI Tian-yu;YANG Yuan-xiao;LIU Hong-yan;WANG Wei;ZHOU Fang;ZHOU Ting;ZHAO Ying-zhong(Oil Crops Research Institute of Chinese Academy of Agricultural Sciences,Key Laboratory of Biology and Genetic Im-provement of Oil Crops,Ministry of Agriculture and Rural Affairs,Wuhan 430062,China)

机构地区：[1]中国农业科学院油料作物研究所,农业农村部油料作物生物学与遗传育种重点实验室,湖北武汉430062

出　　处：《中国油料作物学报》2022年第2期331-343,共13页Chinese Journal of Oil Crop Sciences

基　　金：国家自然科学基金(31701468,31771877);国家特色油料产业技术体系(CARS-14)。

摘　　要：细胞色素P450(CYP450)超家族广泛参与初生和次生代谢物质的生物合成,其中CYP703家族基因参与拟南芥、水稻等的花药发育。本研究利用生物信息学方法,从芝麻基因组中鉴定出303个CYP450基因,发现它们分为9个家族簇(clan),43个基因家族,随机分布于16个连锁群中。CYP703家族是71 Clan的成员之一,为了解芝麻CYP703家族的功能,克隆并分析了其家族基因SiCYP703A2。系统进化分析表明,SiCYP703A2与拟南芥CYP703A2聚为一类,它们的氨基酸序列一致性为77%。表达分析表明,SiCYP703A2在芝麻的花蕾组织中高量表达,同时在花药发育的四分体阶段表达水平较高。拟南芥中超量表达后发现,SiCYP703A2超表达植株荚果变短、花粉活力降低,花药扫描电镜观察显示花粉壁表面出现不规则突起,花粉壁结构异常。此外,荧光定量PCR结果显示,多个花粉壁发育基因在SiCYP703A2超表达植株的花药中异常表达。这些结果表明,超量表达SiCYP703A2会导致转基因拟南芥花粉壁出现缺陷,导致育性降低,推测SiCYP703A2在花粉粒和花粉壁的形成中发挥作用。Cytochrome P450(CYP450s)superfamily is the largest family of enzyme proteins in plants,widely involved in the biosynthesis of primary and secondary metabolites.In this study,303 sesame CYP450 genes were identified,which were randomly distributed in 16 linkage groups,and classified into 9 clans,and 43 gene families.The CYP703 family was one of the members of the 71 Clan.It has been reported that CYP703 genes were involved in anther development in Arabidopsis and rice.Thus the sesam SiCYP703A2 gene,which might be the only member of sesame CYP703 family,was cloned.Phylogenetic analysis of P450 genes in sesame and Arabidopsis showed that SiCYP703A2 and AtCYP703A2 were clustered together,with amino acid sequence identity of 77%.Expression anal‐ysis indicated that SiCYP703A2 was highly expressed in flower buds of sesame,and especially highly expressed at the tetrad stage of anther development.Moreover,SiCYP703A2 in Arabidopsis transgenic plants possed shorter si‐liques and decreased pollen viability.Scanning electron microscopy observation showed that exine of the pollen grains of the overexpression plants appeared irregular protrusions with abnormal pollen wall.In addition,qRT-PCR results showed that several pollen wall development-related genes were abnormally expressed in the anthers of over‐expression plants.These results indicated that overexpression of SiCYP703A2 caused defects in pollen wall and re‐duce fertility of transgenic Arabidopsis.SiCYP703A2 might play a role in formation of pollen grain and pollen wall.

关 键 词：芝麻 细胞色素P450 雄性不育 花粉壁 转基因拟南芥 SiCYP703A2 

分 类 号：S565.4[农业科学—作物学]