title: [Prokaryotic expression, purification of human papillomavirus type 16 E1 protein and preparation of it's antiserum].
abstract: OBJECTIVE
  To express and purify the human papillomavirus type 16 E1 protein in E.coli and prepare the antibody against HPV-16 E1.
  
  
  METHODS
  HPV-16 E1 gene was amplified by PCR and cloned into prokaryotic expression vector pMAL-p2x, and the recombinant plasmid was transformed into E.coil BL-21. We optimize the soluble expression condition of fusion protein by induction with different IPTG concentration and different temperature. The expressed fusion protein was purified by mahose affinity column Chromatography. To prepare the anti-serum, New Zealand white rabbits were immunized with purified HPV-16 E1 protein via hypodermic and volar. Western blot and ELISA analyzed the serum's specificity against HPV-16 E1 and serum titers.
  
  
  RESULTS
  Restriction endonuclease analysis and DNA sequencing showed HPV-16 E1 was cloned into the plasmid pMAL-p2x. Based on the optimization experiments, it concluded that the best soluble expression conditions for the HPV-16 E1 fusion protein involved addition of IPTG to a final concentration of 0.5 mmol/L and then further incubation at 28°C. The purity of the HPV-16 E1 fusion protein was over 95.7% after purification. ELISA and Western blotting showed the titers of the anti-serum were above 1:640 000, and the anti-serum can specifically bind with HPV-16 E1 protein.
  
  
  CONCLUSIONS
  We have ingathered the HPV-16 E1 fusion protein by expressing in E.coli and purifying, and the antibody against HPV-16 E1 was prepared with the fusion protein immunizing New Zealand white rabbits. This work will provide an antigen and detection antibody for further study on the HPV-16 E1 function.
authors.affiliation: College of Life Science and Technology, Xinjiang University, Key Laboratory of Biological Resources and Genetic Engineering, Urumchi 830046, China.
authors.forename: Qian
authors.initials: Q
authors.lastname: Shi
authors.forename: Xiang-qian
authors.initials: XQ
authors.lastname: Hou
authors.forename: Jing-xia
authors.initials: JX
authors.lastname: Guo
authors.forename: Hui
authors.initials: H
authors.lastname: Zhu
authors.forename: Zheng-hai
authors.initials: ZH
authors.lastname: Ma
mesh_terms: D000818:Animals; D000906:Antibodies / Q000276:immunology / Q000302:isolation & purification*; D000918:Antibody Specificity; D003001:Cloning, Molecular / Q000379:methods*; D004926:Escherichia coli; D005822:Genetic Vectors; D006801:Humans; D007106:Immune Sera / Q000276:immunology / Q000302:isolation & purification*; D009856:Oncogene Proteins, Viral / Q000235:genetics*; D010957:Plasmids; D011817:Rabbits; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000276:immunology / Q000302:isolation & purification
chemical_list: D000906:Antibodies; C093478:E1 protein, Human papillomavirus 16; D007106:Immune Sera; D009856:Oncogene Proteins, Viral; D011994:Recombinant Proteins