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Anu Singh-Cundy • Michael L. Cain Discover Biology FIFTH EDITION CHAPTER 16 DNA Technology © 2012 W. W. Norton & Company, Inc.

Genetic Code: 3 things to know 1. Unambiguous: if I show you a codon, there’s no question which amino acid to use next 2. Redundant: most amino acids have more than one codon

Genetic Code: Universal (3) Use same genetic code: Animal, Plant, Bacteria, Fungi, Protist, Archea A gene’s codons have the same meaning even if you put them in a different organism Why we can do this

Artificial Selection: Have done for centuries Old way to manipulate genes • Breed animals/plants • Select any favorable new traits (mutations) • Breed THOSE animals/plants

Manipulating Genes: The Old way People manipulating genes (Selective Breeding) We breed the ones we like, until they are ALL what we want

Other examples http: //www. businessinsider. com/how-fruits-have-evovled-over-time-2014 -10

Belgian Blue Cattle: muscle growth mutation Some cattle grow muscle at a faster rate (mutation turns off genes that regulate muscle growth) Breed THOSE animals until all your cattle have lots of muscle.

People selected for a naturally occurring mutation and only bred the cattle with the allele that increased muscle growth The babies bulk up so fast that they are delivered by cesarean section

DNA techology/Biotechnology A newer way to control genes in organisms • Approx. 40 years (since 1970 s) • Turn a gene on • Turn a gene off • Add a new gene

Manipulating Genes: The new way • Eduardo Kac: Human genes in petunia!!! • Chickens without feathers!!

Nerd Words Restriction Enzymes: How bacteria fight viruses Cut DNA into small pieces cut in specific sequences some leave jagged (“sticky”) ends Restriction Enzymes are the “Scissors” for DNA

Nerd Words DNA Ligase: Used in replication joins fragments enzyme to join DNA fragments used in DNA replication (doubling DNA in interphase) DNA ligase is the “glue” used in DNA Technology joins sugar-phosphate backbone

DNA ligase pastes DNA together

Nerd Words Recombinant DNA: made by combining DNA from different sources Transgenic: DNA from different species (usually unable to breed together) Plasmid: loop of DNA separate from chromosome Plasmids are common In bacteria

Cisgenics adding genes from closely related organisms (often within a species)

Another Nerd Word Intragenics: like cisgenics – i. e. organisms closely related and able to breed – but using technology to make custom combinations of genes

Nerd Words “Amplifying DNA”: making copies of DNA Why? Getting more DNA to run tests Detecting Mutations Crime Scene Paternity Case

Amplifying DNA: the basic idea cut DNA into small pieces Isolate fragment you want (usually a gene) Make many copies of this fragment

Amplifying DNA: 2 methods Making Many Copies of a piece of DNA (gene) 1. With Bacteria put DNA into a bacteria get bacteria to multiply, making more DNA 2. Polymerase Chain Reaction (PCR) Copy DNA outside of a cell uses heating and cooling (more info later)

Nerd Words DNA cloning Putting recombinant DNA into a cell so it will be copied when the cells Often used when studying genetic disorders

Amplifying DNA with Bacteria

Modified Bacteria: Lots of uses • We currently use genetically modified bateria to make insulin blood-clotting proteins for hemophiliacs clot-dissolving enzymes for stroke victims Can use as a vector to transfer genes

Fixing Disease Example: Person has bad copy/copies of gene Give a good copy of gene symptoms reduced/eliminated “Gene Therapy” = Genetic Manipulation to reduce symptoms of a disease

Genomic Editing Scissors: restriction enzymes Smart scissors: zinc finger nucleases & TALENS Precision editing: remove one allele and replace with another example: removing CCR 5 -Delta 32 Making T-cells that are immune to HIV

Polymerase Chain Reaction polymerase chain reaction (PCR) amplifying DNA outside of cells (faster than growing bacteria) Why? Need more DNA than you have to run test When? medical diagnostics, forensics, paternity testing, and paleoanthropology to

Polymerase Chain Reaction Goal: make billions of copies of a targeted sequence of DNA Requires: Template DNA (stuff to be copied) DNA polymerase (special high temp) DNA primers (synthetic) Nucleotides (ACGT) VERY GOOD TEMPERATURE CONTROL

Temperature control in PCR Heat DNA up hot enough to separate hydrogen bonds separates hydrogen bonds of rungs not hot enough to separate sugar-phosphate Cool back down, then repeat

PCR song

DNA Technology Personalized medicine (custom treatments) Gene therapy Genetic Engineering Adding one or more genes Permanently modifies gene/tissue/organism

Genetically Modified Organism (GMO) Created with Genetic Engineering

Genetic Engineering • Transgene: gene introduced into GMO transgenic organism: Another name for a GMO Example: green fluorescent protein (GFP) rabbit, fish, monkey, etc. • Vectors: ways to put new genes into a cell: – Plasmids – Viruses – Gene guns (shoot DNA directly into cell…. usually used for plants)

GMOs: Why make them • Make lots of of a gene product therapeutic or commercial value • Change organism to increase productivity or performance

DNA Fingerprinting • DNA fingerprinting: finding DNA unique to species or individual Forensics (CSI) Paternity Test • Looks at areas of DNA known to have lots of variation

Cloning Plants: Easy • Take a cutting • Grow a new plant • We’ve been doing this for centuries. • Most animals cannot be reproduced this way

Cloning of Animals • Reproductive cloning: making animals that are genetically identical one organism with useful characteristics • Cloning: three-step process 1. removing the nucleus from an egg 2. Fuse this egg with a somatic cell (source of nucleus) 3. Put embryo into surrogate mother

Cloning: The ugly side • High failure rate Example: cloned rat (129 embryos, 3 rats) • May have defects causing Short life span Dolly the sheep (? ? ? ) Pyrenean Ibex (7 minutes, bad lungs) • NOT IDENTICAL!!!! (Just has identical DNA) Brahma Bull

Human Gene Therapy • Gene therapy: fix disorders by altering gene function • Giving people the correct version of a gene • Somatic cell gene therapy – not as controversial • Germ cell gene therapy – much more controversial • RNA interference (RNAi) (the book mentioned it!!!) tricking cells into shutting genes down

Human Gene Therapy • In 1990, Ashanthi De. Silva was the first human to receive gene therapy • Two bad copies of a gene. • White blood cells removed • WBC infected by a virus containing a good copy of gene • WBC put back into body

Human Gene Therapy • RNA interference (RNAi) (the book mentioned it!!!) tricking cells into shutting genes down Make a piece of a gene that looks like double stranded (viral) RNA. Cell destroys the “viral” RNA, and any similar RNA all the m. RNA of gene you want to shut down

Gel Electrophoresis Sorts DNA Fragments by Size • We use electricity to move DNA Sugar-phosphate backbone has charge • DNA moves through gel • Little pieces move faster than big ones Cut with restriction enzymes

Sorting Fragments of DNA • We can sort them even if they are only 1 nucleotide longer • Can determine A, C, G, T • Automated sequencing machines – identify over a million bases per day,

We “borrowed” our tools • Restriction enzymes – used by bacteria to destroy viruses • DNA ligase – used by lots of stuff to join DNA fragments during DNA replication • High temperature DNA polymerase – used by archea that live in hot springs • Plasmids – used by bacteria to transfer genes

Ethics: (Stuff people get worked up about) • Do we have the right to alter DNA of other species? • Are there some species we should not alter? Food? Pets? Primates?

Ethics: (Stuff people get worked up about) • Not just biological issues Social issues Political issues Medicine: “do we have the right…” Farming: “is this sustainable…. ”, “will this harm…. ” “will every farmer be forced to…. ” “do people want to eat…. ”

Monsanto: Love it or Hate it vs

Genetically Engineered Foods Are they dangerous? Should we test? Should we label? some of this is addressed later: always new info Why do we think selectively bred crops are any afer? Lenape potato: solanine

GMOs: judge case by case • Genetically modified crops harmful effects on soil, water, or animals? are they safe? Example: “roundup ready” crops (more poison in soil) vs. BT versions of crops (less poison in soil) Encouraging more poison use is probably a bad idea

Can we control GMOs? Genes can spread (wind blows pollen) people who don’t want to grow it might to so (and get sued by monsanto) What if you can’t eat GMO corn? can you be sure that the corn you buy is free from the things that are bad for you

Legal Regulation of GMOs Reasons people want to ban 1. gene transfer pollen containment via greenhouses 2. food safety 3. superweeds 4. biodiversity 5. “We should know what’s in our food” YES, you should!!!!

Gene transfer Many people worry about wind-born pollen but we have ways to control that

Scientific Consensus: SAFE! Long term studies (yes they exist) discussion on next slide http: //rameznaam. com/2013/04/28/the-evidence-on-gmo-safety/

“There are no long term studies…” Yes there are: multi-year studies fish, cows, mice multi-generational TEN generations in birds three generations in rats No, we cannot prove something is always safe. OTOH, we have not been able to show bad effects

Claim: GMOs cause superweeds More use of pesticides can cause more hardy pests Partially true….

Truth: Weed killers cause superweeds Life sometimes finds a way People use poisons Weeds mutate, some are immune USING POISON MAKES POISON-RESISTANT WEEDS LIKELY Superweeds (develop regardless of GMOs) 70 Poison GMO to resist glyphosate round-up ready atrazine NONE 60 50 40 30 20 Resistant weeds 24 species 64 species http: //www. nature. com/news/case-studiesa-hard-look-at-gm-crops-1. 12907 10 0 roundup atrazine

Biodiversity Human impact always causes winners and losers non-target species are going to be affected especially ones similar to targeted pest Spraying kills everything BT GMO kills things that bite parts of the corn plant

Biodiversity Yes, related species can be killed pollen on wind: monarch effects less than expected plant milkweed elsewhere Net Effect: Pesticides: 30% reduction (Stephen Naranjo) number of species increased

Labeling is coming Obvious, attention grabbing Sort of scary Many options for what it may look like Easily overlooked NO USEFUL INFORMATION!!!

Don’t oppose it, shape it Labeling is coming I support information, not fear Do I trust this company? Which genes? Allows additional research after approval!!! What was the goal?

“But that is scary, too…. ” • “people won’t understand all that…. ” just like normal ingredients labels • If you’re worried that the FDA approved something that it should not have, this is the best way to gather evidence of their mistake • If you think that the approved genetic modifications are safe, this is the best way to gather evidence to support that

Personal Opinion • I care about safety • I want labeling if it is specific enough to allow further research into safety • I don’t want labeling that scares people without allowing further research • I won’t vote for laws that don’t require details (i. e. require labels, and we’ll figure out specifics later)

Clicker Questions CHAPTER 16 DNA Technology

Concept Quiz What is an advantage of restriction enzymes? A. They cut DNA into manageable pieces. B. They cut at a specific sequence and, therefore, give consistent results. C. They leave sticky ends, which can be used in cloning. D. All of the above

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