蜡梅花青素苷合成相关基因CpTT8的克隆和功能分析  

Cloning and Functional Analysis of CpTT8 Related to Anthocyanin Synthesis in Wintersweet(Chimonanthus praecox)

作　　者：张倩 杨楠[1,2] 桑海煜[1] 赵荣 宋晓惜 陈龙清[2] 向林 赵凯歌[1] ZHANG Qian;YANG Nan;SANG Haiyu;ZHAO Rong;SONG Xiaoxi;CHEN Longqing;XIANG Lin;ZHAO Kaige(Key Laboratory of Horticltural Plant Biology,Ministy of Edtcation,College of Horticulture and Forestry Sciences,Huazhong Agricultural University,Wuhan 430070,China;College of Landscape Architecture and Horticulture Sciences,Southwest Forestry University,Kunming 650224,China)

机构地区：[1]华中农业大学园艺林学学院,园艺植物生物学教育部重点实验室,武汉430070 [2]西南林业大学园林园艺学院,昆明650224

出　　处：《园艺学报》2021年第10期1945-1955,共11页Acta Horticulturae Sinica

基　　金：国家自然科学基金项目(31972442)。

摘　　要：以蜡梅(Chimonanthus praecox L.)‘H29’为试材克隆得到1个bHLH类转录因子的全长编码序列,命名为CpTT8。对其生物信息学特征、时空表达特性、基因功能、蛋白互作、蛋白–DNA互作等进行了分析。CpTT8的ORF为2130 bp,编码709个氨基酸,含有典型的bHLH结构域;蜡梅CpTT8与桃PpbHLH3、苹果MdbHLH3同源性最高。实时荧光定量PCR结果表明CpTT8在早期花芽中表达量最高;在根、茎、叶、果实中也有表达,以果实中的表达量最高。在拟南芥Col-0和tt8突变体中超量表达CpTT8有利于花青素苷和原花青素的积累,说明CpTT8是花青素苷合成过程中的正调控因子。酵母双杂交实验和双荧光素酶分析揭示了CpTT8需与MYB转录因子互作才能激活下游结构基因ANS的表达。The full-length coding sequence of a bHLH transcription factor,named CpTT8,was cloned from Chimonanthus praecox‘H29’.Studies related to this transcription factor were carried out,including bioinformatics characteristics,spatiotemporal expression pattern analysis,gene function test,protein interactions,and protein-DNA interaction.Sequence analysis showed that the ORF of CpTT8 consists of 2130 bp encoding 709 amino acids,and CpTT8 contains a bHLH domain.CpTT8 shows the highest sequence similarity to PpbHLH3 from peach and MdbHLH3 from apple.The results of quantitative RT-PCR revealed that the expression level of CpTT8 was highest in flower bud at the early stage of floral development.It was also expressed in roots,stems,leaves,and fruits,with the highest level in fruits.Overexpression of CpTT8 in Arabidopsis Col-0 and tt8 facilitated the accumulation of anthocyanins and proanthocyanidins,indicating that CpTT8 is a critical positive regulatory transcription factor in the anthocyanin biosynthesis pathway.Yeast two-hybrid assay and dual-luciferase analysis illustrated that CpTT8 needs to interact with MYB transcription factors to activate its downstream structural gene ANS.

关 键 词：蜡梅 花青素苷 CpTT8 功能鉴定 互作 

分 类 号：S685.99[农业科学—观赏园艺]