Biotechnology 2007-2008

Biotechnology today Genetic Engineering Electrophoresis Recombinant Technology Polymerase Chain Reaction Transgenics/GMO’s Cloning

Recombinant Technology rDNA is DNA with 2 or more sources Bacteria are used to copy DNA and make desired proteins Identify gene of interest and cut it out insert new gene into a plasmid insert plasmid into bacteria Vector – plasmid carrying new gene into host bacteria can express new gene or make copies of inserted DNA

Restriction Enzymes – Cut the DNA! a.k.a.restriction endonucleases evolved in bacteria to cut up foreign DNA from viruses Molecular scissors that cut between specific bases “Sticky Ends” are where the enzyme cut allow for joining with another piece of DNA cut with the same enzyme

Recombinant tech. Transformation insert recombinant plasmid into bacteria grow recombinant bacteria in agar cultures bacteria make lots of copies of plasmid “cloning” the plasmid production of many copies of inserted gene production of “new” protein transformed phenotype DNA  RNA  protein  trait

insert “gene we want” into plasmid... “glue” together Recombinant Plasmid Plasmids used to insert new genes into bacteria cut DNA gene we want like what? …insulin …HGH …lactase cut plasmid DNA Cut DNA? DNA scissors? ligase insert “gene we want” into plasmid... “glue” together recombinant plasmid

Grow bacteria…make more transformed bacteria gene from other organism + recombinant plasmid vector plasmid grow bacteria harvest (purify) protein

Transgenics – organisms with foreign DNA Genetically modified organisms (GMO) enabling plants to produce new proteins Protect crops from insects: BT corn corn produces a bacterial toxin that kills corn borer (caterpillar pest of corn) Extend growing season: fishberries strawberries with an anti-freezing gene from flounder Improve quality of food: golden rice rice producing vitamin A improves nutritional value For example, a transgenic rice plant has been developed that produces yellow grains containing beta-carotene. Humans use beta-carotene to make vitamin A. Currently, 70% of children under the age of 5 in Southeast Asia are deficient in vitamin A, leading to vision impairment and increased disease rates.

Transformed vertebrates Green with envy?? Jelly fish “GFP” Transformed vertebrates

Many uses of restriction enzymes… Now that we can cut DNA with restriction enzymes… we can cut up DNA from different people… or different organisms… and compare it why? forensics medical diagnostics paternity evolutionary relationships and more…

Copy DNA without plasmids? PCR! Polymerase Chain Reaction method for making many, many copies of a specific segment of DNA ~only need 1 cell of DNA to start It’s a DNA copy machine!

What does 90°C do to our DNA polymerase? PCR process What do you need to do? in tube: DNA, DNA polymerase enzyme, primer, nucleotides denature DNA: heat (90°C) DNA to separate strands anneal DNA: cool to hybridize with primers & build DNA (extension) What does 90°C do to our DNA polymerase?

Can’t we just add those little marshmallows? Comparing cut up DNA How do we compare DNA fragments? separate fragments by size How do we separate DNA fragments? run it through a gelatin agarose made from algae gel electrophoresis DNA jello?? Can’t we just add those little marshmallows?

“swimming through Jello” Gel electrophoresis DNA moves in an electrical field… DNA is negatively charged size of DNA fragment affects how far it travels small pieces travel farther large pieces travel slower & lag behind DNA        – + “swimming through Jello”

Gel Electrophoresis - + DNA & restriction enzyme wells gel longer fragments power source gel shorter fragments + completed gel

Uses: Evolutionary relationships Comparing DNA samples from different organisms to measure evolutionary relationships turtle snake rat squirrel fruitfly – 1 3 2 4 5 1 2 3 4 5 DNA  +

Uses: Medical diagnostic Comparing normal allele to disease allele chromosome with normal allele 1 chromosome with disease-causing allele 2 allele 1 allele 2 – DNA  Example: test for Huntington’s disease +

Uses: Forensics Comparing DNA sample from crime scene with suspects & victim suspects crime scene sample S1 S2 S3 V – DNA  +

Electrophoresis use in forensics Evidence from murder trial Do you think the suspect is guilty? blood sample 1 from crime scene blood sample 2 from crime scene blood sample 3 from crime scene “standard” blood sample from suspect OJ Simpson blood sample from victim 1 N Brown blood sample from victim 2 R Goldman “standard”

Uses: Paternity Who’s the father? – Mom F1 F2 child DNA  +

Clone A genetically identical organism or a group of cells derived from a single cell

Cloning Process The nucleus from a somatic (body) cell is harvested An egg is harvested and the nucleus of the egg is removed The nucleus from the somatic cell is then transferred into the egg An electric shock causes the cell to replicate The embryo is now implanted into the uterus to grow into a new individual

Big Ideas of Biotechnology Genes from virtually any organism can be cloned using plasmid transformation or using PCR. Electrophoresis and restriction enzymes are widely used in DNA analysis. GMO’s and cloned animals and plants can be given beneficial characteristics or make needed products such as pharmaceuticals.