인간의 삶과 역사 속의 미생물

1. 인간의 삶과 역사 속의 미생물 강의자료ppt-8 2010-2학기

2. 미생물은어떤 존재인가?

3. 유전공학(genetic engineering)을 가능하게 한 미생물

4. Genetic engineering (유전공학) The useof in vivo techniques in the isolation, manipulation, alteration, and expression of DNA (or RNA) and in the development of genetically modified organism * Recombinant DNA technology (재조합 DNA 기술) Biotechnology • use of organisms to form useful products

5. Restrictionand modification enzymes • Basic techniques include: - Restriction enzymes (restriction endonuclease, 제한 효소) - Cloning vectors (클로닝매개체) - Molecular cloning (분자클로닝)

6. Restrictionand modification enzymes • Restriction enzymes: • Protect prokaryotes from hostile foreign DNA (e.g., viral genomes) • Widespread among prokaryotes • Rare in eukaryotes • Recognize specific DNA sequences and cut DNA at those sites • Essential for in vitro DNA manipulation

7. Restrictionand modification of DNA

8. Restrictionand modification of DNA • Modification enzymes: protect cell’s DNA for restriction enzymes • Chemically modify nucleotides in restriction recognition sequence • Modification generally consists of methylation(additionof methyl group) of DNA

9. Essentials of molecular cloning • Molecular cloning: isolation and incorporation of a piece of DNA into a vector so it can be replicated and manipulated • Three main steps of gene cloning • Isolation and fragmentation of source DNA • Inserting DNA fragment into cloning vector • Introduction of cloned DNA into host organism

10. Essentials of molecular cloning 1) Isolation and fragmentation of source DNA • Source DNA can be genomic DNA, RNA, or PCR amplified fragments • Genomic DNA must first be restriction digested

11. Essentials of molecular cloning 2) Inserting DNA fragment into cloning vector • Most vectors are derived from plasmids or viruses • DNA is generally inserted in vitro • DNA ligase: enzyme that joins two DNA molecules • Works with sticky or blunt ends

12. Essentials of molecular cloning 3) Introduction of cloned DNA into host organism • Transformation is often used to get recombinant DNA into host • Some cells will contain desired cloned gene, while other cells will have other cloned genes • Gene library: mixture of cells containing a variety of genes (= genomic library) • Shotgun cloning: gene libraries made by cloning random genome fragments

13. Essentials of molecular cloning

14. Plasmids as cloning vectors • Plasmids are natural vectors and have useful properties as cloning vectors • Vector transfer carried out by chemical transformation or electroporation (전기천공)

15. Hosts for molecular cloning (microorganisms)

16. Steps for cloning a gene

17. Restriction enzymes and their recognition sequences

19. Recombinant plasmid construction

20. Genomic library construction and screening by phenotyphic rescue

21. Applications of genetic engineering • In medicine • many useful proteins • gene therapy • In agriculture • use of genetic engineers allows for the direct transfer of desirable traits to agriculturally important animals and plants

22. Social impact of genetic engineering • There is scientific and philosophical concern about • the use of embryonic stem cells in gene therapy. This is currently a major controversial issue • the production of genetically engineered food • the possibility of the production of genetically engineered organisms by bioterrorists • other issues