摘要
生长调控因子(growth-regulating factors,GRF)是植物特有的一类转录因子,该转录因子家族成员数目较多,在植物生长发育过程中发挥着重要作用。以巴西橡胶树优良品种‘热研7-33-97’为材料,通过RT-PCR方法进行HbGRF基因的克隆;利用生物信息学的方法对其蛋白序列、理化性质、基因结构、进化关系进行分析;利用IS-PCR技术和FISH技术对其进行物理定位分析;采用qRT-PCR对橡胶树中HbGRF基因的表达模式进行研究。结果表明:在橡胶树雌花中克隆到3个GRF基因,分别命名为HbGRF1、HbGRF2和HbGRF3,分子量分别为65.663、41.188、52.858 kDa,其编码的蛋白长度分别为609、384、494 aa,均为不稳定蛋白;3个基因均具有GRF完整的特征结构域WRC和QLQ,分别属于3个不同亚族。基因物理定位结果表明,HbGRF1、HbGRF2和HbGRF3基因分别位于橡胶树染色体的第11号长臂、第5号短臂和第9号长臂上,基因到着丝粒的平均百分距离是77.65、42.66和65.27。表达结果显示,橡胶树3个HbGRF基因在茎尖、雌花等生长旺盛的组织中表达量较高,用外源赤霉素和脱落酸处理后,发现表达量呈上升趋势,经过48 h后表达量和初始值基本持平,3个基因在茎尖、雌花中有明显的表达特异性,可能在橡胶树花芽分化及雌花的发育过程中起到了重要作用。该结果为研究橡胶树HbGRF基因的功能及作用机制奠定了理论基础,为橡胶树精准育种提供了分子细胞学依据。
Growth-regulating factor(GRF)is a plant-specific transcription factor.The transcription factor family has a large number of members and plays an important role in plant growth and development.In this study,the excellent clone‘Reyan 7-33-97’was used as the experimental material,and the HbGRF genes were cloned by RT-PCR.The bioinformatics method was used to study its gene length,physicochemical properties,basic structure and evolutionary relationship.The physical localization analysis was carried out by in situ PCR and fluorescence in situ hybridization.The expression pattern of HbGRF genes in rubber trees was analyzed by qRT-PCR.Three GRF genes were cloned into the female flowers of rubber trees,named HbGRF1,HbGRF2 and HbGRF3,respectively.The protein length was 609 aa,384 aa and 494 aa with molecular weight of 65.663 kDa,41.188 kDa and 52.858 kDa,respectively,and they were all unstable protein.All the three members had the complete characteristic domains WRC and QLQ of GRF gene family.Evolutionary analysis showed that the three genes belonged to three different groups.HbGRF1,HbGRF2 and HbGRF3 were located on the long arm of chromosome 11,the short arm of chromosome 5 and the long arm of chromosome 9,and the average percentage distance between the signal site and the centromere was 77.65,42.66 and 65.27,respectively.The expression results showed that the three HbGRF genes in rubber trees were highly expressed in the vigorous growth tissues like stem tips and female flowers.Under the treatment of exogenous GA and ABA,the expression levels of the three genes increased first,and the expression levels at 48 h were basically the same as those under the initial conditions.The three genes had obvious expression specificity in stem tips and female flowers,which may play important roles in flower bud differentiation and female flower development.This study would provide a theoretical basis and materials for further exploration of the function of HbGRF,and provide molecular cytology basis for accurate breeding of rubber trees.
作者
张宇航
潘冉冉
李飞
陶志强
王英
高和琼
庄南生
ZHANG Yuhang;PAN Ranran;LI Fei;TAO Zhiqiang;WANG Ying;GAO Heqiong;ZHUANG Nansheng(College of Tropical Crops,Hainan University,Haikou,Hainan 570228,China;Rubber Research Institute,Chinese Academy of Tropical Agricultural Sciences,Haikou,Hainan 571101,China)
出处
《热带作物学报》
CSCD
北大核心
2020年第9期1723-1732,共10页
Chinese Journal of Tropical Crops
基金
国家自然科学基金项目(No.31760216)
海南省研究生创新科研课题项目(No.Hys2018-13)
国家天然橡胶产业技术体系项目(No.CARS-33-YZ2)。
