Cloning, prokaryotic expression and tissue expression profile of human novel gene HBVDNAPTP1 transactivated by hepatitis B virus DNA polymerase
- VernacularTitle:乙型肝炎病毒DNA聚合酶反式调节人类新基因HBVDNAPTP1的克隆及原核表达与组织表达分析
- Author:
Yongzhi LUN
;
Liying ZHANG
;
Jun CHENG
- Publication Type:Journal Article
- Keywords:
hepatitis B virus;
DNA polymerase;
transactivated protein;
prokaryotic expression;
tissue expression profile
- From:
Medical Journal of Chinese People's Liberation Army
1982;0(01):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To clone the human target gene HBVDNAPTP1 transactivated by hepatitis B virus DNA polymerase obtained by screening with suppression subtractive hybridization (SSH) and bioinformatics techniques. To construct prokaryotic expressive vector of HBVDNAPTP1 gene, induce the expression of recombinant protein in E. coli, and analyze the expression level of HBVDNAPTP1 gene in human tissues. Methods The DNA fragment of HBVDNAPTP1 was amplified by reverse transcription polymerase chain reaction (RT-PCR) taking mRNA from HepG2 cells as the template, and the correct DNA fragment was then inserted into inducible prokaryotic expressive vector pET-32a (+). The competent BL21 (DE3) E. coli was transformed, and then cultured and induced with IPTG. The expressed HBVDNAPTP1 was confirmed with Western blot. UniGene database was used to analyze the chromosome mapping and tissue expression profile of HBVDNAPTP1 gene. Results The DNA fragment of HBVDNAPTP1 was amplified by RT-PCR. HBVDNAPTP1 expressive vector was constructed. After transformation with pET-32a(+)-HBVDNAPTP1 and induction with IPTG, recombinant HBVDNAPTP1 was expressed and confirmed by Western blot. The expression of genomic location of HBVDNAPTP1 gene was low in multiple-tissues with the exception of pituitary gland, tonsil, tongue, thymus, trachea and umbilical cord. Conclusion The recombinant HBVDNAPTP1 gene could be expressed in prokaryotic expression system of E. coli. The chromosome mapping and tissue expression level of HBVDNAPTP1 gene is tentatively conceived.
