Cloning and expression of tilapia Oreochromis aurea gonadotropin-releasing hormone and GnRH associated peptide in Escherichia coli

The cDNA encoding gonadotropin-releasing hormone(GnRH) and GnRH associated peptide(GAP) was amplified from total RNA from tilapia Oreochromis aurea pituitary glands by reverse transcri-ption polymerase chain reaction(RT-PCR) method,and then blasted against other GnRH cDNA sequences in the GenBank.The analysis of the sequence data indicated that the coding region of the cDNA fragment,which encoded 89 amino acid redidues,is about 400 bp in size.The amplified cDNA fragment was cloned into the prokaryotic expression vector,pMAL-c2x,to produce the expression vector pMAL-GnRH.The recombinant plasmid was transformed into Escherichia coli TB1.GnRH-MBP fusion protein was obtained after the addition of IPTG into the growth media.SDS-PAGE analysis revealed that the GnRH-MBP was expressed after induction with IPTG for 4 h.A protein band of 56 000 was observed on SDS-PAGE gel.It is anticipated that the fusion protein will be proved useful for further research.