Cloning Of Mfβ-Hex1 Gene From Medicago Falcata L And Genetically Transformed Into Nicotiana Tabacum L.

In this paper,based on the sequence information of differentially expressed genes in Hulunbeir Medicago falcate with great pastoral traits,a specific primer was designed and the β-glucosaminidase gene of Medicago falcata was cloned by using Polymerase Chain Reaction.The expression characteristics of the gene were studied by reverse transcription-polymerase chain reaction technique and transformed into tobacco by Agrobacterium tumefaciens mediated method which will lay a foundation for further study of the functional characteristics of the gene with the following results:1.A lucerne β-hexosaminidase gene Mfβ-Hex1 was cloned and its cDNA was 786bp which contained an open reading frame as 675bp.It encodes 224 amino acids,mainly composed by glutamic acid,valine and Leucine.The β-Hex gene had a high homology with the β-Hex gene of Tribulus terrestris and Glycine max(Linn.)Merr.respectively.Mfβ-Hex1 gene belongs to the glycoside hydrolase GH20 family without a transmembrane structure while contained a signal peptides and its secondary structure was mainly a-helix.The protein of this gene was unstable and it is located in the cytoplasm.2.The expression of Mfβ-Hex1 gene was related to the grazing tolerance of Medicago falcata.The expression level of Mfβ-Hex1 gene in the plants with higher grazing tolerance was higher than that in the plants with lower grazing tolerance.The expression level of Mfβ-Hex1 in different organs was in the order of leaf>root>stem.3.In this study,pCHF3-Mfβ-Hex1 expression vector was constructed and transgenic tobacco was obtained by Agrobacterium tumefaciens mediated transformation.4.The expression of Mfβ-Hex1 was up-regulated in transgenic tobacco after cutting with a significant level of expression.The expression level of Mfβ-Hex1 gene in transgenic tobacco was leaf>root>stem in turn.