油松PtNF-YC1基因鉴定及其调控球花发育的作用机制研究  

Identification of PtNF-YC1 of Pinus tabuliformis and its molecular mechanism involved in regulation of cone development

作　　者：刘红梅 郑永涛 郭盈添 张晶星 李伟[1] Liu Hongmei;Zheng Yongtao;Guo Yingtian;Zhang Jingxing;Li Wei(School of Biological Sciences and Biotechnology,National Engineering Laboratory of Tree Breeding,Key Laboratory of Genetics and Breeding in Forest Trees and Ornamental Plants of Ministry of Education,Tree and Ornamental Plant Breeding and Biotechnology Laboratory of National Forestry and Grassland Administration,Beijing Forestry University,Beijing 100083,China;National Beipiao Daqingshan Forest Farm,Beipiao 122100,Liaoning,China)

机构地区：[1]北京林业大学生物科学与技术学院,林木育种国家工程实验室,林木花卉遗传育种教育部重点实验室,树木花卉育种生物工程国家林业和草原局重点实验室,北京100083 [2]国有北票市大青山林场,辽宁北票122100

出　　处：《北京林业大学学报》2023年第9期1-8,共8页Journal of Beijing Forestry University

基　　金：国家重点研发计划(2022YFD2200304);国家自然科学基金项目(31860221)。

摘　　要：【目的】针叶树中NF-Y核因子调控球花发育的研究尚未见报导,对油松PtNF-YC1基因克隆、表达特性及功能分析,旨在为针叶树NF-Y基因家族在球花生殖发育中的功能研究提供依据。【方法】(1)利用系统进化树分析油松PtNF-YC1与拟南芥NF-YC亚家族蛋白的亲缘关系;(2)瞬时转化烟草检测PtNF-YC1的亚细胞定位;(3)根据转录组数据分析PtNF-YC1在油松不同组织中的表达特性;(4)PtNF-YC1异源转化拟南芥,分别比较长日照和短日照条件下转基因拟南芥不同株系的开花时间,并对长日照下各株系进行转录组测序,筛选响应PtNF-YC1调控开花的相关基因;(5)通过Y2H和BiFC验证PtNF-YC1与候选蛋白之间互作。【结果】PtNF-YC1基因的开放阅读框为897 bp,编码299个氨基酸,含有典型NF-YC保守结构域,与AtNF-YC3/4/9具有较高同源性。亚细胞定位结果显示,PtNF-YC1定位在细胞核和细胞质。PtNF-YC1在针叶、营养芽、雌雄花芽和根中都能表达,但在雄球花中表达丰度最高。PtNF-YC1异源转化拟南芥可推迟其短日照下开花时间。Y2H和BiFC证明PtNF-YC1与PtCOL5存在相互作用。【结论】PtNF-YC1可调控成花时间,是油松光周期途径诱导球花发育的候选基因。[Objective]The research on the regulation of NF-Y nuclear factor on cone development in conifers has not been reported yet.Through the cloning,expression characteristics and functional analysis of PtNF-YC1 gene of Pinus tabuliformis,it provides a basis for the functional study of conifer NF-Y gene family in the reproductive development of conifers.[Method](1)The relationship between PtNF-YC1 and Arabidopsis thaliana NF-YC subfamily proteins was analyzed by phylogenetic tree.(2)Tobacco was transiently transformed to detect the subcellular localization of PtNF-YC1.(3)The expression characteristics of PtNF-YC1 in different tissues of P.tabuliformis were analyzed based on transcriptome data.(4)PtNFYC1 was heterologously transformed into Arabidopsis thaliana,and the flowering time of different transgenic Arabidopsis thaliana lines under long-day and short-day was compared.The transcriptome sequencing of each transgenic line under long-day was performed to screen the genes related to PtNF-YC1regulating flowering.(5)The protein interaction between PtNF-YC1 and candidate proteins was verified by Y2H and BiFC.[Result]The open reading frame of PtNF-YC1 was 897 bp,which encoded 299 amino acids,had a typical NF-YC conserved domain,and a close relationship with the homologous genes of AtNFYC3/4/9.The subcellular localization showed that PtNF-YC1 was localized in the nucleus and cytoplasm.The analysis of expression patterns in different tissues showed that PtNF-YC1 could be expressed in needles,vegetative buds,male and female cones and roots,but the expression abundance was the highest in male cones and stems.PtNF-YC1 heterologous transformation of Arabidopsis thaliana delayed flowering under short day.It was proved that PtNF-YC1 interacted with PtCOL5 through Y2H and BiFC.[Conclusion]PtNF-YC1 has the function of regulating flowering time and is a candidate gene for inducing cone development through photoperiodic pathway of P.tabuliformis.

关 键 词：油松 PtNF-YC1 光周期调控 球花发育 

分 类 号：S791.254[农业科学—林木遗传育种]