title: [Application of orthogonal analysis to the optimization of HPV16 E2 protein expression].
abstract: This study was aimed to identify pET21b-HPV16E2/BL21(DE3) strain and to optimize the expression of human papillomavirus type 16 (HPV16) E2 protein by orthogonal analysis. Four influence factors on two levels were selected to increase the target protein quantity. The four factors were induction time, induction temperature, inductor concentration and cell density. The quantity of HPV16 E2 protein was used as the evaluation parameter. Induced by IPTG, HPV16 E2 protein was analyzed by SDS-PAGE and Western Blot. Target protein was analyzed by GIS imaging system to quantify the protein level. SPSS13. 0 software was applied to analyze the result. Data showed that the expression strain pET211rHPV16 E2/BL21(DE3) was identified correctly. HPV16 E2 protein expressed mainly at insoluble form. The 42KD protein band was identified by SDS-PAGE and Western blot. Orthogonal test was applied on influence factor analysis and expression optimization successfully. Main influence factors were inductor concentration and induction temperature. The optimimum condition of maximum expression quantity was 37 degrees C, 7h, 1.0 mmol/L IPTG and OD600 1.0. In this experiment, orthogonal test could not only be used to analyze the influential factors and promote the target protein expression, but also be used to provide a better experiment method for molecular biological study.
authors.affiliation: Department of Microbiology, Harbin Medical University, Heilongjiang Key Laboratory of Infection and Immol/Lunity, Pathogenic-biological Key Laboratory, Heilongjiang Higher Education Institutions, Harbin 150081, China.
authors.forename: Qinglong
authors.initials: Q
authors.lastname: Shang
authors.forename: Yanxiu
authors.initials: Y
authors.lastname: Ma
authors.forename: Zhiwei
authors.initials: Z
authors.lastname: Guo
authors.forename: Liqun
authors.initials: L
authors.lastname: Li
authors.forename: Meili
authors.initials: M
authors.lastname: Hao
authors.forename: Yuhui
authors.initials: Y
authors.lastname: Sun
authors.forename: Lanlan
authors.initials: L
authors.lastname: Wei
authors.forename: Hongxi
authors.initials: H
authors.lastname: Gu
mesh_terms: D004268:DNA-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics; D052162:Human papillomavirus 16 / Q000378:metabolism*; D006801:Humans; D009856:Oncogene Proteins, Viral / Q000096:biosynthesis* / Q000235:genetics; D030361:Papillomavirus Infections / Q000821:virology; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics
chemical_list: D004268:DNA-Binding Proteins; C054855:E2 protein, Human papillomavirus type 16; D009856:Oncogene Proteins, Viral; D011994:Recombinant Proteins