Beet M14 Strain Specific Expression Of Genes M14 Of Beta Corolliflora-341 Expression In The Model Plant

Apomictic monosomic addition line of Beta corolliflora Zoss., designated as M14, was obtained from the hybridization and intercross of Beta vulgaris L. and Beta corolliflora Zoss. in sugar beet. M14, which constituted of the normal 18 Beta vulgaris L. chromosomes with the No.9 chromosome of Beta corolliflora Zoss.(VV+1C,2n=19), was found having a chromosome transmission frequency of 96.5% by transmission analysis. M14 served as ideal materials for study of apomixes phenomenon and the apomixes genes, possessing facultative apomixis characteristics that diplospory apomictic reproduction was the main reproductive mode with secondary reproductive mode of sexual reproduction.298 differential ESTs were obtained by the method of SSH and DDRT-PCR, of which one research value EST was selected as candidate ESTs in this study. Full-length cDNA sequences of M14-341 gene was obtained by RACE. Bioinformatics analysis results of M14-341 gene indicated that M14-341 protein contained MADS-box structural domain and K- structural domain . M14-341 gene had similarity of 88% with Spinacia oleracea anthesis transcription factor, which was assessed to be from MIKC family of MADS-box genes.In this research, M14-341 protein was aligned with MIKC family protein of Arabidopsis thaliana by DNAMAN, which indicated that homology of M14-341 protein with AG protein was up to 74%. M14-341 gene was classified as AG genes, belonging to category C genes of"ABCDE"model of MADS-box genes. Eukaryotic expression vector pBI-M14-341 of full-length cDNA sequence of M14-341 gene was constructed. In order to analyze phenotype properties of transformed plants as well as function of M14-341 gene, M14-341 gene was transformed into tobacco and Arabidopsis thaliana respectively utilizing leaf disc transformation and floral dip transformation mediated by Agrobacterium tumefaciens. The result of analysis of PCR , Southern blot, RT-PCR, Northern blot and GUS reaction showed that M14-341 gene was transformed into model plant, and expressed in flowers, stems and leaves in transformed plants. 24 transgenic tobacco plants and 12 transgenic Arabidopsis thaliana plants were obtained. Quantity of M14-341 gene expression was higher in flower of transgenic plant, and lower in leaf and stem. Constitutive expression of M14-341 gene effected florescence and development of floral organ in transgenic tobacco plants, which emerged a series of abnormal phenotypes, such as delaying of florescence, shoaling of petal color, decreasing inflorescence and so on. Degrading of seed setting rate and increasing of fatality appeared in transgenic Arabidopsis thaliana plants.Two-dimensional gel electrophoresis of non-transgenic and transgenic tobacco floral organ total protein were performed. The results showed that expressed protein spots of transgenic tobacco plants are similar to that of non-transgenic tobacco plants. Expressed protein spots ranged from 20 to 100 kD . 8 remarkable differential expression protein spots were detected, including 1 disappeared spot, 1 newly expressed spot, 5 decreasingly expressed spots and 1 increasingly expressed spot.The results of this research indicated that specifically expressed M14-341 gene from M14 has a similar function with AG genes. M14-341 gene plays an important role in differentiation and development of floral organ as well as in regulating florescence.