Genetic technology. Some terminology Genetic engineering –Direct manipulation of genes for...

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?