Studies On Construction Of Plant Expression Vector With BADH Gene Promoted By Rd29A Promoter And Genetic Transformation In Tabacco

To a large extent , successful application of transgenic plant depends on the gene expression level, expression of the foreign genes in transgenic plants was controlled by promoter. The promoter has been a hot spot of plant genetic engineering though expression control of rd29A gene in the experiment, major research as follows:1.A study was designed restriction sites of Hindâ…¢and BamHI, extracted from the plant Arabidopsis genome DNA and cloned of inducible promoter rd29A (907 bp). rd29A promoter region and in Genbank (D13044) has homologous 94.61%,although a small number of mutation or deletion of the base, mutation or deletion cannot influence their activity in the promoter sequence of control of the key sites.The Sequence showed that the promoter has two drought, high salt and low temperature response cis-element (DRE), TATAbox exists in the 474 bp ~ 479bp, in section 597 bp ~ 601bp between a CAATbox (CCAAT), in section 659 bp ~ between 666bp and 714 bp ~ 721bp between each of a drought, high salt and low temperature response(DRE) cis-element, DRE sequence containing nine bases (TACCGACAT).2.Plasmid of pGEMT-rd29A and pBI101.2 of plant expression vector were digested with HindIII and BamHI, and the recovered promoter fragments of 907bp and pBI101.2 line carrier, the recovery of the promoter gene fragments and Linear viscous jointed, through connecting, transforming, and identification of bacteria resistance screening ,it was proved that pBI-rd29A-GUS was constructed successfully .3.Plasmid of pGEMT-rd29A and pBI-BADH of plant expression vector were digested with HindIII and BamHI, and the recovered promoter fragments of 907bp and pBI-BADH line carrier, the recovery of the promoter gene fragments and Linear viscous jointed, through connecting, transforming, and identification of bacteria resistance screening, it was proved that pBI-rd29A-BADH was constructed successfully .4.Plasmid of pBI-rd29A-GUS of plant expression vector was directed EHA105 by freeze-thaw method. It was proved by antibiotic resistance screening and identification of PCR. The transference with the Agrobacterium tumefacien to Nicotiana tabacum were access to transgenic plants...