Cloning, Transformation And Functional Analysis Of Gras Transcription Factor Family Genes In Rice

Using gateway technology, we have cloned44OsGRAS genes from diversetissues such as leaf, spike, root and floral organ in rice (Oryza. sativa L. cv.Nipponbare) and constructed to the reformed plant expression vector pCambia1301including transcription activation expression vector and transcription inhibitionexpression vector. The over-expression of the GRAS genes exhibited differentphenotypes from the other two plant expression vectors, so that we can analysis thefunction of the GRAS gene family members directly.We transformed30OsGRAS genes which constructed into NV transcriptionactivation expression vector to the rice callus by Agrobacterium mediatedtransformation, and selected more than990transgenic lines in T0generation and theseeds of T1generation.We selected3transgenic lines (NV-GRAS04-06, NV-GRAS04-11andNV-GRAS34-22) for the molecular identification according to the field phenotypeof T1generation. PCR detection of the hpt gene showed that93.33%of hygromycin(167mg/L) resistance plants were transgenic plants. PCR analyses using genespecific primers and sequencing results showed that GRAS genes have integratedinto the rice genome. Furthmore, we demonstrated that GRAS genes have expressedat protein level through western blot.T2generations of the two genes OsGRAS34and OsGRAS04, whichconstructed into transcription activation expression vector, showed strong phenotype.The plant height and the stem base width of the transgenic lines at the seeding stageimproved dramatically compared to those of the wild type. The yield of thetransgenic lines raised remarkably compared to the wild type in T1generation in this study. It is suggests that the function of the two genes, OsGRAS34and OsGRAS04,may have concerned with the plant height and the yield. Furthermore, our transgeniclines may provide new genetic resources for rice breeding.