Study On Transformation Of Antisense CCoAOMT Gene In Boehmeria Nivea And Tobacco

The lignin is ,the main chemical composition of plants, the organic compound which quantity is only inferior to the cellulose on the Earth, the evolution product of the plant organization. Lignin plays a vital role in plant's normal growth. However, its existence in plants has negative effects on pulp and paper industry, ramie textile industry and stockbreeding. Therefore, it has been the focus to study lignin biosynthesis and genetic manipulation.Ramie is the China important fiber plants.The lignin content of the raw hemp is 1% ~ 5% ,a few of lignin content is the quite stubborn impurity,and the existence of a few of lignin content still has a tremendous influence to spinning and weaving performance of ramie.What the textile industry use is the ramie's phloem fiber. The raw fiber must be unglued before it is used to spin. During the process of being unglued, the lignin is the impurity which is most difficult to be extracted. It increases not only the cost, but also the pollution. To reach to the ideal effect by conventional breeding or plantation measures always goes in vain.Therefore, reducing lignin content of ramie has the latent prospect through the genetic engineering from the source of the lignin content.In this paper,we transformed the antisence CCoAOMT gene into model tobacco and ramie using antisense RNA techniques. In order to find out whether CCoAOMT of ramie has effectd in the synthesis of lignin and can decrease the lignin content or not.The main results of this study are as follows:(1) We transformed pBSP-antiCCoAOMT-2301 plant expression vector into Agrobacterium tumefaciens LBA4404 by freeze-thaw method,and obtained the works of Agrobacterium strain LBA4404 (pBSP-antiCCoAOMT-2301) by PCR for the plant genetic transformation.(2) In order to verify the availability of the pBSP-antiCCoAOMT-2301 plant expression vector which we had constructed,and measured the possible performance inRamie which we would transformed antisense CCoAOMT cDNA,we transformed pBSP-antiCCoAOMT-2301 into tobacco,and obtained transgenic tobacco by PCR and Southern Blot, and mensurated the lignin content and cellulose content of tobacco which had been transplant in greenhouse about three months. The results showed that the lignin content in the transgenic tobacco had been reduced 16.8%,and the cellulose content in the transgnnic tobacco had been increased 15.2%, compared to non- transgenic tobacco. And the transgenic tobacco can grow well.(3) The genetic transformation system of ramie was optimized in this paper. The medium of induction calla (MS+6-BA3.0mg/L+NAA0.03mg/L+Sugar30g/L+ Agar5g/L), And through the kanamycin resistance experiment of the explants and Ramie sprouts,we confirmed the kanamycin was 50mg/L for the stage of callus induction selection pressure and 40mg/L for root formation selection pressure by experiment.which built a base for plant transformation.(4) Antisense CCoAOMT cDNA had been transformed into ramie. And the transgenic plants were identified by PCR and Southern Blot analysis.