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《Molecular Plant Breeding》 2013-02
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Expression Analysis of Rice Zinc Finger Protein Gene OsC2HC-1 and Response to Salt-alkali and Oxidative Stresses

Zheng Heng Guan Qingjie * Liu Shenkui Key Laboratory of Saline-Akali Vegetation Ecology Restoration in Oil Field, Ministry of Education, Alkali Soil Natural Environmental Science Center, Northeast Forestry University, Harbin, 150040  
Currently, a variety of zinc finger protein genes which involved in abiotic stress response have been discovered in rice, most of them belong to C2H2 type. Study of C2HC type zinc finger proteins was not yet to be reported in rice. In this study, we used RT-PCR technique to clone a known gene (GenBank accession number: AK103785) from cDNA of Nipponbare (Oryza sativa L. spp. japonica). The full length of the coding region was 1 019 bp, which encoded 339 amino acid residuals. We determined that it contained a C2HC zinc finger domain by BLAST, named OsC2HC-1. The coding sequence of OsC2HC-1 was fused with green fluorescent protein (GFP), then transferred OsC2HC-1::GFP into yeast and onion epidermal cells. The results showed that OsC2HC-1 was located in the nucleus. Real-time quantitative PCR exhibited that the transcriptional level of OsC2HC-1 increased significantly in rice roots and leaves induced by the stress of 80 mmol/L NaCl and 30 mmol/L NaHCO3. Prokaryotic expression conditions of pGEX-6p-3::OsC2HC-1 in host strain BL21(DE3) was optimized by a small amount of induction experiments. The results seemed that IPTG concentration had little effect on gene expression, four hour inducing time and 28℃ inducing temperature were the optimal conditions. Fusion protein expressed in Escherichia coli should be insoluble and stable form, which was called inclusion bodies. The purified fusion protein was about 63 kD, which would be a prerequisite for making antibodies. The analysis of resistant experiments after germi- nation presented that overexpression OsC2HC-1-GFP transgenic Arabidopsis significantly enhanced abiotic stress tolerances under 35S promoted, including 125 mmol/L NaCl, 1 mmol/L NaHCO3 and 3 mmol/L NaHCO3, indicating that OsC2HC-1 should be strong responsive to the salt-alkali stress as well as oxidative stress.
【Fund】: 中央高校基本科研业务费专项资金(DL11CA05);; 国家林业局948项目(2008429);; 中国博士后基金(2012M-520695)共同资助
【CateGory Index】: S511
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