The. Osrack1 Of Response To Drought And Salt Stress Function And Its Fusion Protein Expression And Purification Of Rice Seed Germination

Drought is one of the most severe natural disasters, drought has caused serious disruption to the growth and development of rice growing period significantly slowed, resulting in crop failures. Therefore, improvement of water utilization rate and adaptation to drought of rice has become key problems which are urgently needed to resolve. The previous study showed gene rack1 mutant was drought tolerance in Arabidopsis thaliana. The Atrack1 mutant was screened that can tolerate osmosis stress from Arabidopsis. Amino acid sequences of two genes (NC008394 and NC008398) expression in the Oryza sativa were found high homology with AtRACK1 of Arabidopsis thaliana from NCBI. The two proteins of genes expression are 70% identity and 80% similarity in the protein level AtRACK1. We deduce that OsRACK1 may regulate rice osmosis stress. This study will focus on the function of OsRACK1 under drought stress. Over-expressing plantsof RACK1, anti-sense plants of RACK1 and control were choosed as mantierials. The main results are as follows:1. In order to identify T2 and T3 generations of transgenic plants, GUS dyeing, southern blotting and RQ-PCR were utilized to know the situantion of gene expression. Truebred transgenic plants were obtained.2. Bourgeon experiment were performed for the seed of T3. The content of soluble sugar, soluble protein, SOD and MDA were measured after seeds were treated with various contentration of PEG6000. The over-expressing seeds sprouted better than others. The content of soluble sugar and SOD is higher than anti-sense and control, but the content of protein and MDA is lowest.3. The genes encoding RACK1 from Oryza santiiva genome were cloned into plasmids pGEX-6P-1, constructed the expression plasmid pGEX-6P-1/OsRACK1.We use the BL21 expression system to express Oryza santiiva RACK1. The optimal concentration of IPTG, an inducer of the recombinant plasmids is 0.1mM for the expression plasmids. The optimal expressing temperature is 24oC. Furthermore, about 50%RACK1 protein is expressed in the soluble fraction. Through a series of purification procedures, including differential centrifugation, affinity chromatography and SDS-PAGE, the purity of RACK1 are 90% at least.