摘要
[目的]本文旨在研究不结球白菜Br CDF1基因的功能,验证其与CO启动子之间的调控关系,以及其在植物抽薹开花中所起的作用。[方法]以不结球白菜‘苏州青’为材料,采用酵母单杂交技术,验证Br CDF1转录因子与CO启动子是否能够结合;构建过表达载体p Early Gate101-Br CDF1,利用农杆菌介导法将过表达载体导入拟南芥中,筛选阳性苗,同时对阳性苗进行Western blot检测,观察转基因植株的表型变化;采用RT-q PCR技术,检测Br CDF1、CO、FT基因在转基因和野生型植株中不同时间点的相对表达量。[结果]酵母单杂交和β-半乳糖苷酶试验分析结果显示,Br CDF1转录因子和CO启动子之间具有较高的结合活性。Western blot检测结果表明:转基因拟南芥植株中的Br CDF1蛋白成功表达。RT-q PCR结果表明,Br CDF1基因在转基因植株中的相对表达量比野生型植株高;CO、FT基因在转基因植株中的相对表达量比野生型低,同时Br CDF1过表达植株较野生型开花晚。[结论]Br CDF1转录因子能与CO启动子结合,从而抑制CO、FT基因的表达,进而影响不结球白菜光周期开花途径。
[Objectives]The paper aimed to study the function of BrCDF 1 gene in non-heading Chinese cabbage,and to verify its regulatory relationship with CO promoter and its role in bolting and flowering of plants.[Methods]The yeast single hybridization technique was used to verify whether BrCDF1 transcription factor could bind to CO promoter by using the non-heading cabbage‘Suzhouqing’.Then the over-expression vector pEarlyGate101-BrCDF 1 was constructed and introduced into Arabidopsis thaliana by Agrobacterium-mediated method.The positive seedlings were detected by Western blot.The phenotypes of transgenic plants were observed.The expression of BrCDF 1,CO and FT genes in transgenic and wild type plants at different time points was detected by RT-qPCR.[Results]The results of yeast single hybridization andβ-galactosidase(ONPG)assay showed that BrCDF1 transcription factor and CO promoter had strong binding activity.Western blot analysis showed that the expression of BrCDF1 protein in transgenic Arabidopsis plants was successfully expressed.The results of RT-qPCR indicated that the relative expression level of BrCDF 1 gene in transgenic plants was higher than that in wild type plants.The relative expression level of CO and FT genes in transgenic plants was lower than that in wild type plants,and BrCDF 1 overexpression plants showed later flowering than wild type plants.[Conclusions]The BrCDF 1 transcription factor could bind to the CO promoter,thereby inhibiting the expression of CO and FT genes and thus affecting the photoperiodic pathway regulating flowering in non-heading Chinese cabbage.
作者
李林
陈丽
吴小婷
邵帅旭
李英
侯喜林
刘同坤
LI Lin;CHEN Li;WU Xiaoting;SHAO Shuaixu;LI Ying;HOU Xilin;LIU Tongkun(State Key Laboratory of Crop Genetics and Germplasm Enhancement/Key Laboratory of Biology and Germplasm Enhancement of Horticultural Crop in East China,Ministry of Agriculture and Rural Affairs/College of Horticulture,Nanjing Agricultural University,Nanjing 210095,China)
出处
《南京农业大学学报》
CAS
CSCD
北大核心
2018年第5期832-838,共7页
Journal of Nanjing Agricultural University
基金
江苏省自然科学基金项目(BK20171374)
中央高校基本业务费专项资金(Y0201700179)
关键词
不结球白菜
BrCDF1
酵母单杂交
功能特性
non-heading Chinese cabbage
BrCDF 1
yeast single hybrid
functional characterization
