Biotechnology Use of Natural Biological Systems to Produce a Product or Provide a Desired Process

Recombinant DNA Technology A set of techniques used to produce large quantities of a gene and its product Recombinant DNA = DNA produced by joining segments of DNA from different sources eg. To produce human insulin, scientists have combined bacterial DNA + human DNA

Tools for Producing Recombinant DNA Restriction enzymes: enzymes that cleave the DNA double helix at specific nucleotide sequences

Use of the Restriction Enzyme Bam H1 5’— G G A T C C — 3’ 3’— C C T A G G — 5’ Results in sticky end 5’— G G A T C C — 3’ 3’— C C T A G G — 5’

Tools for Producing Recombinant DNA Vector: carrier of DNA; can be virus or plasmid Plasmid: extrachromosomal, independently replicating, small circular DNA molecule

Producing Recombinant DNA restriction enzyme Mix together Add DNA Ligase Many recombinant DNA molecules are produced, each with a different piece of source DNA Treat source DNA with restriction enzyme Treat plasmid DNA with same enzyme restriction enzyme Transform bacterial cells Each bacterial cell carries a different recombinant plasmid

Tools for Producing Recombinant DNA Probe: sequence of DNA that is complementary to the gene of interest; Used to locate a copy of the gene by hybridization AGCTTAGCGAT TCGAATCGCTA Denature DNA by heating AGCTTAGCGAT TCGAATCGCTA AATCGC Add Probe Probe Binds to gene

Using the Probe to Find the Gene of Interest

Biotechnological Methods: PCR http://www. dnalc PCR = Polymerase Chain Reaction Amplifies a specific region in the DNA Used for identification, especially if the amount of DNA is small Uses repeated cycles of heating to denature DNA and cooling to synthesize new DNA Involves the use of ---Taq polymerase (a DNA polymerase that withstands heat) ---primers to begin synthesis

DNA Fingerprinting with PCR

DNA Fingerprinting for Paternity Testing

DNA Fingerprinting in Forensics 1 2 3 C S R C I E M N E E 4 5 6 7 Suspects Suspects

DNA sequence for gene from cress plant Biotechnological Methods DNA Sequencing: Determining the Order of Nucleotides Arrows show differences in DNA sequences related to inherited breast cancer DNA sequence for gene from cress plant

Human Genome Project An effort to determine the sequences of all human genes Genomics: study of DNA sequences Proteomics: study of proteins produced in a specific organism or cell type Bioinformatics: using computer databases to store and analyze sequence information

Applications of Biotechnology Transgenic Bacteria Transgenic: organism that contains a gene from another species in all of its cells Transgenic bacteria that produce Human Insulin Human growth factor tissue plasminogen activator (t-PA) hepatitis B vaccine

Applications of Biotechnology Transgenic Plants Bt Corn: Produces its own Pesticide Roundup Ready Soybeans are resistant to herbicide “Golden” rice with beta-carotene and extra iron

Human gene product will be released in milk Applications of Biotechnology Transgenic Animals Make Human Gene Products Human gene product will be released in milk

Same type of mouse “rescued” by blocking action of human gene Applications of Biotechnology Transgenic Animals as Models of Human Diseases Transgenic Mouse with defective human hormone gene interfering with growth Same type of mouse “rescued” by blocking action of human gene

Ex vivo: Gene is introduced into cells outside the body Andrew Gobea Ex vivo: Gene is introduced into cells outside the body Gene Therapy for SCID

Aerosol spray delivers gene Cystic Fibrosis In vivo Gene Therapy Aerosol spray delivers gene Injection of gene Blindness: Leber’s Congenital Amaurosis (LCA)