大豆转录因子GmC2H2基因转化拟南芥效果分析  

Effects Analysis of Transcription Factor GmC2H2 in Transgenic Arabidopsis

作　　者：单曙光[1] 于国红[1] 徐娜[1] 程宪国[1] 

机构地区：[1]中国农业科学院农业资源与农业区划研究所农业部植物营养与肥料重点实验室,北京100081

出　　处：《生物技术通报》2011年第12期75-81,共7页Biotechnology Bulletin

基　　金：国家转基因重大专项(2008ZX08002-005)

摘　　要：GmC2H2转录因子基因是本实验室获得的一个编码172个氨基酸携带516 bp核苷酸的转录因子,属于经典C2H2型锌指蛋白。通过构建植物表达载体GmC2H2-pCAMBIA1304,借助优化的Floral-dip法转化模式植物拟南芥,经潮霉素Hygromycine(45-50 mg/L)抗性筛选获得转基因拟南芥植株。GUS组织染色分析表明,GmC2H2基因在生长12 d的转基因拟南芥幼苗中,表达部位主要集中在根部。对转基因拟南芥进行了低温(1℃)和脱落酸(200μmol/L)胁迫处理,测定其生理生化指标,通过real-time qPCR确定目的基因在转基因拟南芥中的表达情况。结果表明,携带GmC2H2目的基因的转基因拟南芥中脯氨酸和可溶性糖水平要高于野生型植株,而丙二醛水平要低于野生型,在抗逆性方面明显优于野生型拟南芥植株;并且胁迫处理下的转基因拟南芥中GmC2H2基因的表达量要高于未胁迫处理的转基因植株,说明GmC2H2基因的表达受低温和ABA的诱导,初步明确了该转录因子基因的功能。A transcription factor named GmC2H2 is consisted of encoding 172 amino acids with an open reading frame of 516 bp length,and belonging to the classical C2H2 zinc finger protein family.In this study,GmC2H2-pCAMBIA1304,a high efficient expression vector,was constructed and transferred into wild type Arabidopsis using floral-dip method.13 regenerated plant were selected in T0 generation by Hygromycine（45-50 mg/L）,and 10 positive seedlings were obtained by PCR and GUS detecting.The positive ration was 77%.Histochemical staining analysis of the report gene GUS showed that GmC2H2 was expressed in the roots from 12-days-old of transgenic Arabidopsis thaliana.In this study,to determine expression of GmC2H2,the transgenic Arabidopsis plants were treated with 1℃ and ABA,respectively.To determine the effect of stress resistance and expression patterns of GmC2H2 gene,physiological and biochemical indexes and real-time qPCR were determined.The increased levels of free proline and soluble sugars were observed in transgenic Arabidopsis with containing the gene GmC2H2 compared to wild-type plants under salt stress,while the MDA has a lower level related to wild type plants.Accumulation of GmC2H2 mRNA is higher in transgenic Arabidopsis after treatment with low temperature at 1℃ and ABA.This shows that GmC2H2 was induced by low temperature and ABA,and initially defined the function of the transcription factor genes.

关 键 词：GmC2H2 锌指蛋白 转基因拟南芥 GUS检测 REAL-TIME qPCR 

分 类 号：S565.1[农业科学—作物学] Q943.2[生物学—植物学]