- To serve as an important medium for the publication of original research in the field of medical science and health research, thus filling gaps in health knowledge for effective utilization of research findings
- To impart current medical knowledge and updated scientific information obtained from research to health professionals for better and appropriate health care management
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Archives 2009
Myanmar Health Sciences Research Journal
Volume 21, Number 1
TITLE: Cloning of the hepatitis B surface antigen gene (1): extraction and tranformation of the recombinant HBsAG gene containing plasmid into the E. coli DH5 alpha cells.
AUTHOR: Win Aung; Yeon Hee Kim; Hyunah-Kim; Suk Hoon Ha; Khin Pyone Kyi
SOURCE: Myanmar Health Sciences Research Journal. 2009; 21(1): 56-61
ABSTRACT: The majority of currently available recombinant hepatitis B (HB) vaccines in the markethave been produced by using appropriate expression plasmids in Saccharomyces cerevisiae and Hansenula polymorpha yeast cells as host systems. Recently, attempts weremade to use Pichia pastoris as an alternative and more productive host for cloning of yeast expression plasmid carrying the hepatitis B surface antigen (HBsAg) structural gene for production of recombinant HB vaccines in the future. DNA cloning involves separating a specific gene or DNA segment from a larger chromosome, attaching it to a small molecule of carrier DNA, and then replicating this modified DNA thousands or millions of times through both the increase in cell number and the creation of multiple copies of the cloned DNA in each cell, followed by expression of desired protein. In this study, the HBsAg gene was first extracted and purified from chromosomal DNA of H. polymorpha transformant cells. It was then ligated with pGEM-T vector followed by transformation into the competent E. coli DH5 alpha cells which had already beenprepared in our laboratory. The results from each and every steps were confirmed by direct PCR identification, restriction enzyme analysis followed by agarose gel electrophoresis determination and DNA sequencing analysis. Thenucleotide sequences of the HBsAg gene, extracted and purified from the final transformant E. coliwere found to be totally identical to that of the HBsAg gene which had been initially extracted and purified from H. polymorpha cell. Research works on amplification and ligation ofthe HBsAg gene and yeast plasmid in E. coli DH5 alpha cells for further transformation of the recombinant HBsAg gene containing plasmid into the P. pastoris yeast cells are in progress at the Research and Development Centre of Pharmaceuticals, Institute of Science and Technology, CJ Corporation, Ichon City, Republic of Korea.
SUBJECT HEADINGS: Cloning, Organism. Hepatitis B Surface Antigens. Plasmids. Recombination, Genetic. Escherichia coli.
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