Expression Profile Analysis Of OsCPKs & Research Of Hvppc2 Transgenic Rice

Calcium-dependent protin kinases(CDPKs) is a kind of Ca²⁺ receptor,playing important roles in calcium-meadiated signal transduction involed in plant growth and development,responses to abiotic stresses,defenses to pathogen,ionic transportation in channels,in the research,we identified 31 rice CDPK genes by searching the rice genome sequences databases,and discussed the evolution of OsCPKs based on the phylogenetic tree analysis and intron analysis.Using the rice gene expression profile database(CREP: http://crep.ncpgr.cn) in our lab,we carried on the expression profile analysis of OsCPKs, and tested candidate genes by Real-time PCR.The main results are as follows:1.Thirty-one rice CDPKs genes were identified in rice genome through sequence camparing with conserved domain of plant CDPKs genes,the putative amino acid sequences have typical CDPKs domains,such as Ser/Thr kinase domain,autoinhibits region,CaM-like domain.Based on the structure analysis,all CDPKs contain 4 EF-hands except OsCPK30.In addition,21 OsCPKs were found to contain myristoylation sites at their termini,28 CDPKs were predicted to contain the palmitoylation sites.2.Most OsCPK genes had five to seven introns,while OsCPK4,OsCPK18 and OsCPK30 had 10-11 introns,all three genes belong to Groupâ…£.Combined the result that both two CDPKs genes from Chlamydomonas belong to Groupâ…£,the members in Groupâ…£maybe the oldest during the evolution.3.The expression of this gene family was analyzed using the Affymetrix rice genome array in three rice cultivars:Minghui 63,Zhenshan 97,and their hybrid Shanyou 63 independently.Twenty-seven tissues sampled throughout the entire rice life-span were studied,along with three hormone treatments(GA₃,NAA and KT),applied to the seedling at the trefoil stage.All 31 OsCPK genes had their corresponding probes set on the DNA chip.All genes were found to be expressed in at least one of the experimental stages studied and revealed diverse expression patterns.4.Eight genes,OsCPK2,OsCPK11,OsCPK14,OsCPK22,OsCPK25,OsCPK26, OsCPK27 and OsCPK29 were found dominantly expressed in the panicle and the stamen, and five genes,OsCPK6,OsCPK7,OsCPK12,OsCPK23 and OsCPK31 were up-regulated in the endosperm stage.Showed that they may involved with seed development5.The OsCPK genes were also found to be regulated in rice seedlings subjected to different hormone treatment conditions,as OsCPK5 was up-regulated under the thress hormone treatment.The increasing of population and the decreasing of fields are threatening the crop safety.Enhancing the crop photosynthesis to increase the crop yield is important to agricultural research.It is hot to clone the C₄ genes from C₄ plant,and to create C₄ transgenic rice.The identification of single-celled C₄ plant makes it come to true.In the research,we cloned the C₄ PEPC gene from Hydrilla verticillata,and developed the transgenie rice.1.The PEPC activities of Hvppc2 transgenic rice are between 1-4 folds compared the wild type;the activities of NADP-ME and PPDK haven't changed in the transgenic rice.2.Some Hvppc2 transgenic rice showed high net photosynthesis rate under high light condition,and no change in dark respiration.3.The proline content was increased in Hvppc2 transgenic rice under PEG-6000 modulated drought condition.But no changes soluble sugar content,MDA content and SOD activity.So Hvppc2 transgenic rice may tolerant to drought.4.In filed test,no agricultural characteristic changes in Hvppc2 transgenic rice except the plant height,which became shorter than wild type.