Molecular Clonging And Expressing Of Prothoracicotropic Hormone And Juvenile Hormone Esterase From Heliothis Viriplaca Hufnagel

Insect neuropeptides play key roles in regulation of metabolism, development, reproduction, rhythm, diapause, behavior and learring, and it is important to demonstrate the gene transcriptional regulation of insect neuropeptides. Prothoracicotropic hormone (PTTH) plays a central role in controlling growth, molting, and metamorphosis in insects by stimulating the prothoracic glands to synthesize and release the molting hormone, eedysone.The juvenile hormones (JHs) represent a family of acyclic sesquiterpenoids that are, with a single known exception, unique to the class Insecta. They plays a central role in controlling growth, metamorphosis, and reproduction in insects. Juvenile hormone (JH) metabolism enzymes, including JH esterase, JH epoxide hydrolase and JH diol kinase, catalyze JH to metabolize. By the action of these enzymes, JH is metabolized to JH acid, JH diol, JH acid diol, and JH diol phosphate. The activity of these enzymes can be controlled to influence the growth of insects, and the genetically engineered bacteria and transgenic plants could be constructed by using those genes to against insect pests.In this study, the mRNA was isolated from the insect prepupal stage. The cDNA sequence of Heliothis viriplaca Hufnagel PTTH and JHE were cloned by RT-PCR and rapid amplification of cDNA ends (RACE)The Heliothis viriplaca PTTH cDNA,823base pairs in length, contained an open reading frame of681base pairs, coding for a polypeptide of262amino acid residues with a predicted molecular weight of26.3kDa and pI8.15. The deduced amino acid sequence is composed of a signal sequence, a precursor domain and a mature hormone including seven conserved cysteine residues and hydrophobic regions.The cDNA sequence has been deposited in GenBank with accession No. JF731347. One putative N-glycosylation sites and one O-glycosylation site were found in the amino acid sequence. PTTH was cloned into the expression vector pET21b and expressed in E. coli (BL21) host cell. Induced by IPTG, the protein was expressed and detected in the cells.The Heliothis viriplaca JHE cDNA,3106base pairs in length, contained an open reading frame of1746base pairs, coding for a polypeptide of581amino acid residues with a predicted molecular weight of63.9kDa and pI5.11. The cDNA sequence has been deposited in GenBank with accession No. JQ901384. The juvenile hormone esterase gene of Heliothis viriplaca was cloned into the expression vector pET21b and expressed in E. coli(BL21)host cell.