Molecular Cloning And Sequence Analysis Of SNARE Gene From Japanese Sea Perch

Membrane trafficking and exocytosis of the eukaryotic cell appear to be regulated through the action of set of proteins called SNAREs. To reveal the function of the SNAREs in teleostean, gene specific PCR primers were designed according to the conserved region of cDNA sequences presented at GenBank database. Reverse transcription PCR (RT-PCR ) was conducted and cDNA partial sequences of core SNAREs proteins SNAP-25, VAMP2 and Syntaxinl were successfully amplified from Japanese sea perch. Based on information from RT-PCR sequencing results, more primers were redesigned and used subsequently in Rapid Amplification of cDNA Ends (RACE) to acquire more sequence information of cDNA. Whole sequence of SNAP-25, eighty percent of VAMP2 and half of Syntaxinl gene were finally cloned from Japanese sea perch. Sequence analysis of SNAP-25 gene including homologous comparison, protein primary structure identification and secondary structure prediction were performed by using biological web servers and related software. The characteristics of SNAP-25 protein structure, relationship with other SNARE core proteins were analyzed based on the RT-PCR sequencing results and deduced functions were discussed.