Changes of DHN1 expression and subcellular distribution in A.delicisoa cells under osmotic stress  

作　　者：邱全胜 王泽宙 蔡起贵 姜荣锡 

机构地区：College of Life Sciences, Beijing Normal University, Beijing 100875, China[1] Institute of Botany, Chinese Academy of Sciences, Beijing 100093, China[2]

出　　处：《Science China(Life Sciences)》2002年第1期1-9,共10页中国科学（生命科学英文版）

基　　金：This work was supported by the Major State Basic Research Development Program of the People's Republic of China, the National Natural Science Foundation of China (Grant No. 39770077) ; the Science and Technology Program of the Ministry of Education of

摘　　要：The changes of DHN1 expression and subcellular distribution in A. delicisoa cells under osmotic stress were studied by using GFP as a reporter molecule. Through creating the Xba I and BamH I restriction sites at the ends of dhn1 by PCR, the expression vector for the fusion protein DHN1-mGFP4 was constructed by cloning dhn1 into plasmid pBIN-35SmGFP4. Then the DHN1-mGFP4 expression vector was transformed into A. delicisoa suspension cells by microprojectile bombardment method. Bright green fluorescence of GFP which shows the high-level expression of DHN1-mGFP4 was visualized after culture for 10 h. However, the green fluorescence was only located within the nucleus. By increasing the culture medium osmotic potential, the green fluorescence was visualized in the cytoplasm (mainly around the plasma membranes). The generation of GFP fluorescence in the cytoplasm was also promoted by increasing the medium osmotic potential. Moreover, GFP green fluorescence was abolished by protein synthesis inhibitor dicyclohexylcarbodiimid, indicating that the cytoplasmic DHN1 was newly synthesized under osmotic stress. Furthermore, ABA promoted the presence of green fluorescence in the cytoplasm, and the GFP fluorescence was visualized within a shorter time under a higher osmotic potential.The changes of DHN1 expression and subcellular distribution in A. delicisoa cells under osmotic stress were studied by using GFP as a reporter molecule. Through creating the Xba I and BamH I restriction sites at the ends of dhn1 by PCR, the expression vector for the fusion protein DHN1-mGFP4 was constructed by cloning dhn1 into plasmid pBIN-35SmGFP4. Then the DHN1-mGFP4 expression vector was transformed into A. delicisoa suspension cells by micropro-jectile bombardment method. Bright green fluorescence of GFP which shows the high-level expression of DHN1-mGFP4 was visualized after culture for 10 h. However, the green fluorescence was only located within the nucleus. By increasing the culture medium osmotic potential, the green fluorescence was visualized in the cytoplasm (mainly around the plasma membranes). The generation of GFP fluorescence in the cytoplasm was also promoted by increasing the medium osmotic potential. Moreover, GFP green fluorescence was abolished by protein synthesis inhibitor dicyclohexylcarbodiimid, indicating that the cytoplasmic DHN1 was newly synthesized under osmotic stress. Furthermore, ABA promoted the presence of green fluorescence in the cytoplasm, and the GFP fluorescence was visualized within a shorter time under a higher osmotic potential.

关 键 词：A. delicisoa suspension cells  OSMOTIC stress  DEHYDRIN DHN1  green fluorescent protein (GFP)  transient expression  SUBCELLULAR distribution  ABA. 

分 类 号：Q24[生物学—细胞生物学]