In Silico Cloning And Verification Of AmTpi CDNA From Apis Mellifera

Triosephosphate isomeraser(TPI) is expressed by nuclear gene and participate in glucose catabolism in cytoplasm which translate phosphodihydroxyacetone into glyceraldehyde 3-phosphate. TPI is a dimer structured by the same two subunits which are formed by 246-255 amino acid residues, molecular weight is about 27.5 Kda. Various souces of TPI are quite conservative by judging the primary structure, tertiary structure and mode of action are analogical. The mutation of Tpi will retard decomposition of phosphodihydroxyacetone; accelerate glyconeogenesis; and restrain glucose catabolism, that causes neurological diseasse. At present, the Tpi has been cloned from some kind of species, such as human, rat, drosophila, mosquito, yellow mealworm etc., the anagenesis of Tpi is high conservation.A novel Apis mellifera,triosephosphate isomerase (Tpi) gene is cloned by blasting its EST database and PCR test,and its sequence has been deposited in the GenBank (Accession No:EF493864). Its full length is 1 768 bp, and has a complete open reading fragment (ORF) and encodes 247 Aa protein. The sequence analysis of Tpi's molecular evolution and the result can inflect the relationships of Tpi evolution and classification. cDNA sequencing analysis reveals that the exon fragments are consistent with AmTpi. In the process of insilico cloning, we found there was a piece of sequence had not been sequenced in genome of Apis mellifera, and then it was supplemented by sequencing.