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Genetic technology
Some terminology
• Genetic engineering– Direct manipulation of genes for practical
purposes
• Biotechnology– Manipulation of organisms and their
components for practical purposes
• Recombinant DNA – DNA from 2 or more different organisms
Applications of Genetic engineering
• Manufacturing proteins
• Transgenic agriculture
• Forensics
GE tools
• DNA cloning
• Restriction enzymes
• Plasmids
• Gel electrophoresis
• DNA sequencing
Restriction enzymes
• Restriction Enzymes– Protection against viruses
• Restriction site– Palindrome, enzyme specific
Recombinant DNA
• Sticky ends are complimentary
• Form Hydrogen bonds
• DNA ligase solidifies
Sugar-phosphate
backbone
Cloning Vector
• Correct recombination isn’t very efficient• Screen bacteria for
individuals that carry
the desired plasmid/gene• Antibiotic resistance• Nucleic acid probe
– Complimentary to – desired sequence– Fluorescent tag
DNA cloning
• Foreign DNA is inserted into a plasmid
• Recombinant bacterium
• Bacterium copies
plasmid when it
reproduces
• Bacteria can
produce protein
Genomic Library
• Produce a bacterial colony for each gene
• Grow colonies,
select desired
individual using
nucleic acid probe
Amplifying DNA: PCR
• Heat denatures DNA
• Add primers that
appear on either
side of desired gene
• Only DNA between
primers is copied
Restriction Fragment Analysis
• Mutations can alter restriction sites, alter where DNA is chopped
• Gel electrophoresis– Sorts fragments by size– Agarose Gel
Mapping Genomes
• Linkage maps based on recombination– Map units
• Physical map based on DNA structure– Base pairs
DNA sequencing
• Dideoxy nucleotide
ends chain
• Fluoresces in laser
• Separate
segments by size
Genomics
• Study entire genomes
• Mine sequence for information
• Study interaction and expression of genes
Medical Applications
• Diagnosis
• Human Gene Therapy
• Pharmaceuticals
Gene Therapy
• Genetic Diseases are incurable
• Provide dividing cells with correct allele
• Some cells begin to produce correct protein
• Bone marrow cells divide throughout lifetime
Considerations/ risks
• Linked to cancer
• Reaction to virus
• Can’t always control expression of correct gene
• “Corrections” to germ line cells?
Pharmaceuticals
• Human proteins– Insulin– Human Growth Hormone
• Vaccines– Attenuated viruses– Recombinant viruses
• Virus treatment– Drugs that mimic cell receptors
DNA fingerprinting
• Non-coding regions of genome differ more than coding regions
• Use Gel electrophoresis to separate fragments
• Stain DNA with probes
Forensic Evidence
• Blood and tissue type can only exclude suspects, can’t prove links
• Restriction sites within non-coding DNA are unique
• Use PCR to amplify DNA
Agricultural Applications
• “Pharm” animals
• Transgenic crops
• Environmental Cleanup
Transgenic animals
• Contain genes from other species– Muscle development– Human proteins
Transgenic Plants
• Pesticide– Bt corn, soybeans, cotton
• Herbicide– Roundup Ready
• Nutrients– Golden Rice
• Pharm plants– Hepatitis vaccine– Toothe decay
Environmental Cleanup
• Bioengineered bacteria metabolize waste products– Heavy metals– Oil – Plastic– Chlorine
Concerns?
• New pathogens?
• Outcompete native species?
• “Escaped” genes
• Economic impact of GM crops?
• Privacy?
• Profit?