The Characters And Functions Of Helicoverpa Armigera Molt-Regulating Transcription Factor-Hormone Receptor 3

Helicoverpa armigera is a kind of great harmful insect, which brings huge ecnomy loss to our country and the world every year. Tranditional control approachs are not suit with the new demands for its low effect and severe environment pollution. Biological control by restraining its ecdysis is a better way to control this pest. This thus requires very detailed understanding the ecdysis mechanism of H. armigera. As we know, the whole ecdysis process involves numerous genes and other assistant factors, which is a set of complicated molting cascade. To understand this process, we should first choose the candidate genes and then research the genes and the factors involved in the cascade. By this way, we can grasp the ecdysis mechanism step by step. In this experiment, the candidate gene I choosed for studing is HHR3.Molt-Regulating Transcription Factor (Hormone Receptor 3, HHR3) belongs to an orphan nuclear recepors. It is the classical expression product of molt-relating ealy gene cluster, and is characteristic of regulating molt-relating gene expression by binding to unique DNA sequence. HHR3 is the vital factor in the molting cascade. It is a member of hormone receptor (HR3) family and regulates the ecdysis process of H. armigera. The full length of HHR3 gene is 1668 bp, which encodes a protein with 556 amino acids. Its molecular weight of HHR3 is 61 kDa, and the fusion protein, GST-HHR3 expressed in E.coli, is about 94 kDa.H. armigera is choosed as the experimental material and HHR3 is choosed as the candidate gene in this study. First, GST-HHR3 fusion protein expressed in E.coli is extracted. H. armigera genome DNA was subsequently digested by Rsa I and linked with a adaptor I. Polymerase chain reaction (PCR) is then used to improve the quantity of DNA fragments. The PCR products and GST-HHR3 protein were combined with each other. By use of the antibody against HHR3, the protein that bond DNA is precipitated. After three times of panning, phenel and chloroform is used to extract DNA. This DNA is thus used as a template for further PCR amplification. The control experiment is performed by the same way but replaces GST-HHR3 with PBS. In this control experiment, none of DNA is obtained.To investigate the structure of HHR3 gene, several degenerated primers based on the known HHR3 gene sequence are used to study the introns. One intron sequence is clarified using genome DNA as a templet. By comparing with HHR3 gene, the location and size of intron is analyzed.T7 Select Phage Display system is used to investigat cofactors of HHR3. The H.armigera epidermis T7 phage library and GST-HHR3 protein are used for this study. Through panning a binding protein is abtained.