Expression Of WIL-15 And Function Analysis

Interleukin 15 (IL-15) is a member of the four-helix bundle cytokine family and has Tcell growth factor activity. IL-15 plays a unique role in both innate and adaptive immunecell homeostasis, particularly for the development of NK cells and CD8+ memory cells. Itmay be useful for stimulation of specific immune responses in chronic viral infection suchas hepatitis B virus infection. The woodchuck model is an informative animal model forstudies on hepadnavirus infection and therapeutic interventions. In this study the wlL-15protein were expressed in prokaryocyte and insect cells by using different expressionsystems. Purified protein was acted on mouse splenoeytes to show the effect of purifiedwlL-15 protien on promoting the proliferation of activated mouse splenocytes.Hepatitis C virus (HCV) is a major etiologic agent of posttransfusion and sporadiccommunity-acquired non-A, non-B hepatitis.Recently, serveral research groups reported theHCV in vitro culture systems, which highly enhance the basic research of HCV. Tring to setup infectious clone of HCV Chinese strain, we isolated and cloned the HCV-Hubei straincDNA genome.This thesis includes sthree chapters.In chapter one, a brief introduction of discovery of HCV and HBV, the molecularcharacteristics of them, the genotype of the HCV and HBV, the diagnose method to test thegenotype and the therapy way to patient infected with HCV and HBV. We also introducethe function of wIL-15 and the protein expression system.In chapter two, the pMAL and Bac-to-Bac system had been used to express thewIL-15 protein in prokaryocyte and insect cells, respectively. 55 kDa protein had purifiedBy ion exchange chromatography, and digested by Factor Xa to divide MBP-tag. TheMBP-wlL-15 protein and wIL-15 protein were acted on mouse splenocytes but did't showthe effect of promoting the proliferation of activated mouse splenocytes. Eukaryotic expressed protein was 18 kDa, bigger than theoretic value because of the post-translationmodification.Chapter three discribes cloning of HCV-Hubei genome. Sequence analysis showed thegenome was 9405 nt in length (GenBank accession number: EF638081), belonged to HCV1b subtype, and most homoeologous to HCV-Shanghai. So far the full cDNA genome wascloned into 3 plasmids,was 3 kb, 4.9 kb and 2 kb in length. The genome included a 8952 ntopen-reading form, encoded a polypeptide of 2864 aa.