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

Diagnosis Probes –Build a probe to hybridize with HIV genome Restriction Enzymes –Mutations disrupt restriction sites –Changes fingerprint Sequencing –Build primers that isolate disease gene –Amplify using PCR –Identify fragment using electrophoresis

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?