东亚飞蝗海藻糖酶基因的克隆与表达分析-微生物学专业论文.docxVIP

ABSTRACT Trehalose (a-D-glucopyranosyl-a-D-glucopyranoside), a disaccharide composed of two glucose molecules linked by an a-1-1-bond, is a multi-functional non-reducing sugar in a diverse range of organisms including bacteria, fungi, plants and all kinds of insects. Trehalose exists in insects as the predominant sugar (80-90% of total sugar) in the haemolymph of insects and plays an important role in the energy metabolism. It provides a carbohydrate store and protects proteins and cellular membranes from environmental stresses, such as desiccation, dehydration, excessive heat, freezing and oxidation. Trehalase is the specificity which hydrolyze one molecule of trehalose into two molecules of glucose. In view of the fact that trehalase plays an important role in energy metabolism of biological organisms, in recent years, many biololgists and chemists are devoted to the research and development of new agricultural chemicals, which based on inhibitting the activity of trehalase, blocking trohalose metabolism achieving the insect disinfestation and the bacteriostasis goals. Trehalase plays an important role in trehalose metabolism. There have been many reports about trehalases in insects, the cDNA sequences encoding trehalase have also been cloned. The trehalase genes in locusts, a global agricultural pest, have not been studied. The complete cDNA of a Trehalase gene of Locusta migratoria manilensis (Meyen) was cloned by means of RT-PCR，3’-RACE and 5’-RACE. (GenBank accession number: FJ795020). The result showed that the cDNA was 1953bp in length and contained an open reading frame (ORF) of 1686bp, which encoded 561 amino acids with a calculated molecular weight of 65579.82 Daltons and pI of 6.66. Signal peptide and transmembrane analysis showed that this protein belongs to soluble trehalase. The amino acids showed a high similarity with trehalase of other insects(41-48%). Semi-quantitative RT-PCR indicated that the LmTre was highly expressed in the active tissues in