Sugar Beet M14 Strain Bvm14 Rab, Bvm14 By Mads Box Gene Expression Analysis

Sugar beet apomictic monosomic addition line M14carrying an alienchromosome9of Beta corolliflora Zoss. was secelted through the hybridization andintercross posterity of Beta vulgaris L. which are excellent genetic breeding materialsand Beta corolliflora Zoss., this chromosome provided M14with many excellentcharacteristics of Beta corolliflora Zoss. in the background of full set chromosomes ofBeta vulgaris L., therefore, researching the specifically expressed genes in M14hasgreat value and significance.The Rab family, an important molecular swith controlling vesicule trafficking, canbind and hydrolyze GTP to regulate the action of protein synthesis, endocytosis,degradation and secretion, moreover, stress-resistant processes in plants. MADS-boxgene family mainly affect flowering periods and floral organ development in plants, inaddition, participate the vegetative growth period, it is a kind of transcription factorwhich can regulate transcriptions of other genes and plays an important role in thecourse of growth and development of plants.Previously, the cDNA full lengths of BvM14-Rab gene and BvM14-MADS boxgene have been already obtained in the use of DDRT-PCR、SSH�'�RACE technology;prokaryotic expression vector named pET-28a-BvM14-Rab have been constructedsuccessfully, finding out the optimal condition fit for BvM14-Rab protein expressingaccording to orthogonal experiment; the transgenic tobaccos which are T₂generationtransformed with BvM14-Rab have been obtained and the expression detection inmolecular level have been operated, never in protein level.Based on the previous research, the transgenic tobaccos transformed withBvM14-Rab have been detected in protein expression level and the specific expressionanalysis of BvM14-Rab gene and BvM14-MADS box gene in different tissues of M14 lines have been in progess, it could provide foundation for further research of genes.In this study, recombinant BvM14-Rab was expressed in inclusion body form inEscherichia coli, purified in Ni+affinity chromatography method and eluted in EluteBuffer with imidazole concentration of300mmol/L. it was demonstrated thatBvM14-Rab fusion protein has9antigenic determinants, the antiserum of immunizedNew Zealand rabbits was prepared for Western blot that can confirm the successfulexpression of BvM14-Rab gene in transgenic tobaccos.It was known in the results of specific expression analysis of BvM14-Rab geneand BvM14-MADS box gene in different tissues of M14lines, BvM14-Rab expressedsignificantly in stems and also expressed in other tissues in similar degree;BvM14-MADS box gene expressed significantly in flowers, pistils and stamens, but italso expressed in roots, stems and leaves in different degree.