The Establishment Of Agropyron Mongolicum Keng Regeneration System And Functional Analysis Of Dreb Genes

Agropyron mongolicum Keng not only has a cold, drought and other outstanding features, and contains excellent resistance gene. In this study,the regeneration system was Establishment of Agropyron mongolicum Keng, the DREB gene was separation and the functional was studied.The results as follows:1. Establishment of Regeneration System of Agropyron mongolicum KengAgropyron mongolicum Keng Shoot tip as the most suitable explants for callus induction,the optimal medium of callus induction was MS+2.4-D1.5mg/L+6-BA0.1mg/L,the induction rate of84.9%.The optimal medium of differentiation was MS+6-BA2.0mg/L+NAA0.5mg/L with the differentiation rate of53.2%. The medium of rooting was MS+NAA0.5mg/L with rooting rate of100%.2. Genetic transformation of Agropyron mongolicum Keng DREB geneThe Agropyron mongolicum Keng DREB gene inserted between the vector pCAMBIA2300-35S-OCS promoter and terminator,The plant Expression Vector was constructed of DREB Gene of PCAMBIA2300-MDREB.The plant expression vector of PCAMBIA2300-MDREB was transformed into Agrobacterium tumefaciens LB4404, infecting the tobacco leaf discs,30transgenic tobacco plants were selection by resistance regeneration culture.15positive plants ware determined by PCR.7plants appeared PCR-Southern hybrid spectrum belt, and six transgenic tobacco by RT-PCR validation results show that the Agropyron mongolicum Keng DREB gene expression at the transcriptional level.3. The resistant function analysis of transgenic tobacco1) Transgenic tobacco leaves wilting degree lower than the wild-type tobacco under drought stress and the Turgor-Pressure in Leaves recovery faster than wild-type tobacco after watering.2) Transgenic tobacco membrane relative conductivity is lower than wild-type tobacco and the SOD activity, free prolinecontent are higher than wild-type under drought stress.3) The transgenic plants are obviously dwarf.