미생물 재조합 보고서

소개

미생물 재조합에 관한 보고서입니다.
방선균을 사용하여 올리고마이신을 저해하고 유용물질인 아버멕틴을 생산하는 방법에 대해 기술하였습니다.
영문 초록도 포함하였으며 거의 논문 형식에 따라 썼으며
자세한 설명으로 미생물에서의 재조합 과정의 실험적인 면을 보고싶으시면 참고할만 합니다.

목차

Abstract
Introduction
Materials
Methods
Discussion
Reference

본문내용

Abstract
Streptomyces is a Gram-positive, aerobic soil bacterium, has growth pattern similar with fungus. Streptomyces is well known for their ability to synthesize an enormous variety of antibiotics as secondary metabolites. Among them, S. avermitilils produces avermectins, a group of antiparasitic agents used in human and veterinary medicine. However, S. avermitilis also produces oligomycin, which is a potential toxic inhibitor of oxidative phosphorylation in mammalian cells. Therefore, we decided that disruption oligomycin synthetase gene to prevent co-production of oligomycin in S. avermitilis. To create plasmid for disruption, the smallest gene of oligomycin synthease gene cluster was obtained by PCR from S. avermitilis. And also to create plasmid for disruption, the gene of avermectin A synthase, AveD. Then, apramycin resistance gene was inserted in oligomycin synthase gene and avermectin A synthase gene for selection. After transformation of this plasmid, oligomycin synthase gene(olmA1) and avermectin A synthase gene(AveD) in the chromosome was displaced on the plasmid via homologous recombination. And next, to create plasmid for over expression, the gene of avermectin synthase aglycon. Used pQE30 vector to gene transfer.
As a result of this way, we obtain mutants(olmA1 and AveD) and transgenic cell that no longer not makes the toxic oligomycin and unusable avermectin but more amount of most useful avermectin B.

참고자료

· [1] Hotson, I.K., The avermectins: A new family of antiparasitic agents. J S Afr Vet Assoc, 1982.
· [2] Omura, S., et al., Genome sequence of an industrial microorganism Streptomyces avermitilis: deducing the ability of producing secondary metabolites. Proc Natl Acad Sci USA, 2001.
· [3] Bierman, M., et al., Plasmid cloning vectors for the conjugal transfer of DNA from Escherichia coli to Sterptomyces spp. gene, 1992.
· [4] Hwang, Y.S., et al., Cloning and analysis of a DNA fragment stimulating avemectin production in various Stertomyces avermitilis strains. Appl Environ Microbiol, 2003.
· [5] Hopwood, D.A., et al., Genetic manipulation of streptomyces: A laboratory manual. The John Innes Foundation, F. Crow & Sons Ltd., Norwich, England, 1985.
· [6] Cole, et al., Method for the seperation of antibiotic macrolides. United States Patent 4,160,861, 1979.