by Ms P H Giri Department of Microbiology Deogiri College Aurangabad BSc T Y Semester VI Paper No XIX Recombinant DNA Technology Ms Priyanka H Giri Unit 2 Vector ID: 1043909
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1. Topic –Vector:Presented byMs. P. H. GiriDepartment of MicrobiologyDeogiri College, Aurangabad
2. B.Sc. T. Y. Semester VIPaper No. XIXRecombinant DNA TechnologyMs. Priyanka H. Giri
3. Unit 2
4. Vector:Vectors are DNA molecules, which can carry a foreign DNA fragment to be cloned.These are self replicating in an appropriate host cell.Most important vectors are plasmids, bacteriophages, cosmids and artificial chromosome vectors.
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8. Typesofvectors:
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16. Types of vectors:pBR322pUC18Bacteriophage vectors (improved λ vector)CosmidsYAC (Yeast Artificial Chromosome)
17. 1. pBR322
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19. pBR322 is a plasmid and was one of the first widely used E. coli cloning vectors. Created in 1977 in the laboratory of Herbert Boyer at the University of California, San Francisco, it was named after Francisco Bolivar Zapata, the postdoctoral researcher and Raymond L. Rodriguez. The p stands for "plasmid," and BR for "Bolivar" and "Rodriguez." pBR322 is 4361 base pairs in length and has two antibiotic resistance genes – the gene bla encoding the ampicillin resistance (AmpR) protein, and the gene tetA encoding the tetracycline resistance (TetR) protein.
20. It contains the origin of replication of pMB1, and the rop gene, which encodes a restrictor of plasmid copy number. The plasmid has unique restriction sites for more than forty restriction enzymes. Eleven of these forty sites lie within the TetR gene. There are two sites for restriction enzymes HindIII and ClaI within the promoter of the TetR gene. There are six key restriction sites inside the AmpR gene.
21. The circular sequence is numbered such that 0 is the middle of the unique EcoRI site and the count increases through the TetR gene. The AmpR gene is penicillin beta-lactamase. Promoters P1 and P3 are for the beta-lactamase gene. P3 is the natural promoter, and P1 is artificially created by the ligation of two different DNA fragments to create pBR322. P2 is in the same region as P1, but it is on the opposite strand and initiates transcription in the direction of the tetracycline resistance gene.