Cloning And Function Analysis Of Rice OsICK1 Gene

Cell division is one of the most important processes of plant growth and development. In eukaryotic species, the cyclin-dependent kinase (CDK) is the central component of the regulatory machinery controlling the progression of the cell cycle.The cyclin-dependent kinases (CDKs) are specific serine/threonine kinases that control progression through the cell cycle in all eukaryotes. The activity of CDKs and their substrate specificity depend on their association with cyclins and interaction with other proteins. The cyclin-CDK complexes regulate the cell cycle through the phosphorylation of down-stream substrates at key points of cell-cycle progression, i. e. the transitions from the G1 phase to the S phase and from G2 to mitosis. The cyclin-dependent kinase inhibitors(ICKs) plays a negative role in regulating the cell cycle of eukaryotic organisms including plants. CDK inhibitors are a group of low-molecular-weight proteins that have the ability to bind to cyclin-CDK complexes, thus affecting the activity of CDKs. So ICK can effect the processes of plant growth and development. And as a cell-cycle regulatory, We have been paying much attention to the fuctions of plant ICK.In our research, We have obtained a rice ICK-like gene, and designated it as OsICK1. Then We constructed the plant expression vector of OsICKl gene. Through Agrobaterium mediation transformation system, We have successfully made OsICKl gene fused into the rice gene, and now analysis the functions of OsICKl gene. PCR, Southern dot blotting, Southern blotting and GUS detection have demonstrated that OsICKl gene have been successfully fused into the rice (zhong hua 11) genome. Results of Real-time PCR have showed that transcription level of OsICKl gene in five plants is higher than control one. overexpression of OsICKl gene can effect plant development. All the work was summarized as belows:(1) Rice OsICKl gene obtainedBy RT-PCR , We obtained a ICK gene from rice 9311 and named OsICKl. Sequenced results show that full length of OsICKl gene is 585bp, encoding 194 amino acid.(2) Overexpression vector constructionThrough vector pBPF , we obtained the gene segment of P35s~\- OsICKl + Tnos. Then the gene segment was inserted into plant expression vector pCAMBIA1301, and named 1301+OsICKl.(3) Transformed plant obtainedWe obtained 24 OsICKl gene transformed plants in 27 regenerated plants. PCR, Southern dot blotting, Southern blotting and GUS gene expression detections have demonstrated that OsICKl gene have been successfully fused into the rice (zhong hua A) genome.(4) OsICKl gene transcription level detectionBy Real-time PCR, We obtained 5 plants their OsICKl gene transcription level are higher than others.(5) anti-OsICKl polyclonic antibody preparationWe have obtained the anti-OsICKl polyclonic antibody. It can be used to analysis OsICKl expression level in transgenic plant.