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DNA Cloning

DNA Cloning. Cloning. a line of genetically identical cells or individuals derived from a single ancestor produces many copies of a piece of DNA uses a little fraction as gene of interest cultivates a large amount for studying functions. Functions. Biological determine the DNA sequence

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DNA Cloning

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  1. DNA Cloning

  2. Cloning • a line of genetically identical cells or individuals derived from a single ancestor • produces many copies of a piece of DNA • uses a little fraction as gene of interest • cultivates a large amount for studying functions

  3. Functions • Biological • determine the DNA sequence • ultimate information • regulation • Pharmaceutical • manufacture medicines • Clinical

  4. How it Works • Combine gene of interest and bacterial plasmid • Recombinant DNA • DNA from two or more different sources that have been joined together to form a single molecule • Amplification and identification

  5. Zoom In!!! • Insertion, the most critical step • made possible by restriction endonucleases(aka. restriction enzymes) • recognize specific DNA sequences called restriction sites • cut the DNA at specific locations • Restriction fragments

  6. Restriction Enzyme • defend against viral attack by breaking down the DNA molecules of infecting viruses • cleave the sugar-phosphate backbones of DNA to produce sticky ends • short single-stranded regions • form hydrogen bonds with complementary sticky ends on any other DNA molecules cut with the same enzyme • kind of like glue when you fix something

  7. Cloning Vectors • DNA molecules into which a DNA fragment can be inserted to form a recombinant DNA molecule for the purpose of cloning • plasmid cloning vectors • contains 2 unique genes: ampR gene and lacZ+ gene

  8. The Two Genes • ampR gene • encodes an enzyme that breaks down the antibiotic ampicillin • lacZ+ gene • encodes ß-galactosidase (hydrolyzes the sugar lactorse and synthetic substrates)

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