Genetic Engineering and Recombinant DNA Technology Diversity of life is based on the differences in genes (DNA) 1 gene = 1 protein = 1 function? If we change the DNA (genes), then it would be possible to change the function of the cell, and ultimately the organism

Genetic Engineering and Recombinant DNA Technology Recombinant DNA-isolating and combining DNA of one organism with the DNA of a different organism Human Insulin gene  bacteria Genetic Engineering-modifying gene(s) to benefit or cure an organism Agriculture  crops, animals Gene therapy: when cloned genes are used to modify humans, to control genetic diseases  sickle cell, hypercholesterolemia Ice cream anyone?

GloFish

Cloning Genes Cloning-production of genetically identical DNA To clone a gene, must combine human DNA with another organism’s DNA  recombinant DNA Put gene into a plasmid  extra bacterial chromosome that can accept foreign DNA

Molecular Tools Used For Recombinant DNA How to combine the DNA from 2 different organisms? DNA must be cut and put back together What kind of molecule could cut DNA and put it back together again?

Molecular Tools Used For Recombinant DNA: Restriction Enzymes Bacterial enzymes that cut DNA at specific DNA sequences Leaves single stranded tails at the end of the DNA fragments  ”sticky ends” DNA fragments cut by same enzyme will come together and form a 2x strand DNA fragment; sticky ends are complementary DNA ligase closes the “nicks” in the DNA

Molecular Tools Used For Recombinant DNA: Polymerase Chain Reaction (PCR) Multiple copying of a specific DNA sequence Requires: primers Heat insensitive DNA polymerase from Thermus aquaticus (Taq) Used when only have small amounts of DNA and need to do many different tests; genetic identification Cycle 30 = 5.37 x 10 8

Genetic Fingerprinting Every person has specific DNA sequences that are unique; like fingerprints During a criminal investigation, those are the sequences that are studied using restriction enzymes, PCR and gel electrophoresis Gel electrophoresis-isolation of DNA using electrical currents and a gel matrix made of sugars

Genetic Fingerprinting: Gel Electrophoresis DNA has (-) charge, it will run towards the bottom of gel that has (+) charge Fragments separate based on size. Match fragment pattern with DNA found at scene. Take DNA found at crime scene Restriction enzymes, STRs, PCR Gel electrophoresis Compare DNA fragments  name suspect

DNA Sequencing Allows for the actual nucleotide sequence to be known Automated, done by machines Used to “solve” all the genes in humans  Human Genome Project Is there an advantage to knowing the entire gene sequence of an organism? Functional genomics and comparative genomics

Applications of Genetic Engineering In agriculture: Strawberries that resist frost rice with beta carotene tomatoes that last longer pest resistant corn (95% modified) Larger animals  more meat, milk In environment: Bacteria that clean up oil/toxic spills In medicine: Insulin, growth hormone, interferon, taxol, relaxin, erythropoietin

Genetic Engineering Problems: Is it safe for human consumption? Allergies? If organisms get out into natural setting, impact environmental balance? Bigger animals  more waste, competition Hybrids: herbicide resistance gene in corn passed to a weed The “Enviropig” has been genetically modified in such a manner that its urine and feces contain almost 65 percent less phosphorus than usual.Enviropig

Gene Therapy  Introducing the normal gene into humans with disease  We can make the genes through rDNA, but how do we get them inside to every cell?  Ex vivo gene therapy uses modified viruses to get the new gene inside cells  SCID, familial hypercholesterolemia  In vivo gene therapy uses direct injection or inhalation of normal/healthy gene to defective cells/organs  How do we prevent an immune reaction?