Notes:

Write down ONLY the things in

RED!!!

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Genetic Engineering• Means making changes to DNA

in order to change the way living things work.

• Creates new crops and farm animals.• Makes bacteria that can make medicines.• Can grow human body parts.• Can prevent genetic diseases, change humans.

Altering Organisms isn’t NEW, we’ve been doing it for 1000’s of years…

It’s called - Selective Breeding.

Selective Breeding

For Example: The Labradoodle• Look at the following dogs - crossing a Poodle and a

Labrador results in a ‘Labradoodle’• What features has the Labradoodle inherited from the

Labrador?• What features has the Labradoodle inherited from the

Poodle?

Poodle Labrador Labradoodle

+

+

The simple addition, deletion, or manipulation of a single trait in an organism to create a desired change.

-Major tool is recombinant DNA.

-Recombinant DNA (rDNA) - DNA joined to other unrelated foreign DNA.

-also called gene splicing.-tiny segments of a gene are taken out and replaced.

Transgenic Organisms:

Are organisms that have been altered by genetic engineering.

• Genetic material changed by other than random natural breeding.

• Gene transfer - moving a gene from one organism to another.

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Examples of Genetically Modified organisms:

Enviropig

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Venomous cabbage

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Web-spinning Goat

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Fast Growing Salmon

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Less Flatulent Cows

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Banana Vaccine

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Cancer Fighting Eggs

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How does it work?

In cross pollination (think Mendel’s peas), we are combining two traits to get a mixture of results

In genetic engineering, a single gene, a half page recipe in the 52-thousand-page set of recipe books, can direct the plant to make new traits or remove them

This diagram shows how one type of GM food, a strawberry that resists frost damage is made. The flounder is a fish that live in icy seas. It has a gene that stops it from freezing to death. Strawberries are soft fruits that can easily be damaged by frost.

1. The flounder’s antifreeze gene is copied and inserted into a small ring of DNA taken from a bacteria cell.

2. The DNA ring containing the flounder gene is put into a second bacterium.

3. This second bacterium is used to infect the strawberry cell. The flounder’s antifreeze gene enters the strawberry’s DNA. Strawberry cell

with Antifreeze gene4. The new GM

strawberry cell is grown into a GM strawberry plant which can be bred many times.

Thanks to the new gene, GM strawberries make a protein which helps them resist frost. They don’t contain any other fish genes, and do not taste or smell of fish.

Wonder what they used to make this one blue? – A different gene from another organism.

Pros of Genetic Engineering:• Crops with enhanced characteristics

• Better taste and quality • Less time to ripen.• More nutrients, more food, and stress tolerance • Improved resistance to disease, pests, and herbicides • New products and growing techniques

• Enhanced Animals • Increased resistance, productivity, hardiness, and feed efficiency • Better yields of meat, eggs, and milk • Improved animal health

• Environment (helps clean up and keep it clean)• "Friendly" bioherbicides and bioinsecticides • Conservation of soil, water, and energy • Better natural waste management • More efficient processing

• Helps with increased food demands• More food for growing populations

Cons of Genetic EngineeringSafety • Potential human health impact: allergens, transfer of antibiotic resistance

markers, unknown effects • Potential environmental impact: unintended transfer of transgenes through

cross-pollination, loss of flora and fauna biodiversity Access and Intellectual Property • Domination of world food production by a few companies • Increasing dependence on Industralized nations by developing countries Ethics • Violation of natural organisms' intrinsic values • Tampering with nature by mixing genes among species • Objections to consuming animal genes in plants and vice versa • Stress for animal Labeling • Not mandatory in some countries (e.g., U. States) • Mixing GM crops with non-GM confounds labeling attempts

Crops can be given extra genes for new and useful characteristics. They are genetically modified (GM).What characteristics

might be useful in crops? pest resistance

frost resistance

herbicide resistance

drought resistance longer shelf life

disease resistance

GM crops

Potatoes can be genetically modified so they are toxic to pests, such as the Colorado beetle.

The gene for a powerful bacterial toxin is added to the potato plant.If the beetle tries to eat the potato plant, it is killed by the toxin.

Pest-resistant crops

Crops can be genetically modified so they are resistant to adverse environmental conditions.

For example, lettuces could be genetically modified to be resistant to frost.

GM lettuce

non-GM

lettuceWhy are some people against the development and use of GM crops?

Frost-resistant crops

Rice can be genetically modified to make beta-carotene, a substance that is converted into vitamin A in the body.

Plants with extra vitamins

The GM rice is called ‘Golden Rice’ and is being developed to help fight vitamin A deficiency and blindness in developing countries.

The colour of the rice is an indication of how much more beta-carotene it contains.

The First Clone!Her name was Dolly.

Now cats may have more than nine lives. The company that funded the first successful cloning of a domestic cat, has gone commercial. You can clone your own kitty. Your cost? U.S. $50,000 each.

"Cc," the first-ever cloned cat shown here at seven weeks old with Allie, her

surrogate mother.

The cat was cloned by transplanting DNA from Rainbow, a female three-colored tortoiseshell (or calico) cat into an egg cell whose nucleus had been removed, and then implanting this embryo into Allie, the surrogate mother. "CC's coat color suggests that she is a clone, and a genetic match between CC and the donor mother confirms this," the researchers say.

How does it work?

In cross pollination (think Mendel’s peas), we are combining two traits to get a mixture of results

In genetic engineering, a single gene, a half page recipe in the 52-thousand-page set of recipe books, can direct the plant to make new traits or remove them

Recombinant DNA• The ability to combine

the DNA of one organism with the DNA of another organism.

• Recombinant DNA technology was first used in the 1970’s with bacteria.

1982• Humulin® is approved for the treatment of

diabetes.

Step 1 Isolation of Foreign DNA

• Involves finding the gene you want to sequence’

• Then cutting it out of the chromosome (DNA) with restriction enzymes (sticky ends).

Restriction Enzyme Example

• TaqI

T CGAAGC T

• Cuts between T and C• Leaves sticky end CG

Recombinant DNA

Step 2 Insertion of DNA into Bacterial Plasmid

• Cut the plasmid (DNA) with restriction enzyme.

• Insert gene (DNA) of interest into the plasmid, forming recombinant DNA.

Step 3 Transformation• Insert recombinant plasmid

into bacteria. • Bacteria produced with the

recombinant DNA expresses the gene of interest.

Transformed BacteriaWith Fluorescence

Genetically Modifying Organismshttp://www.hhmi.org/biointeractiv

e/genetic-engineering

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Golden Rice Debatehttps://www.youtube.com/watch?

v=Ayv_EYi43E8

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Transformation of Bacteria

Transformed Bacterium

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BINARY FISSION

• Asexual Reproduction– Copies chromosome– Attach to cell’s plasma

membrane• DOUBLING THEIR

NUMBERS EVERY 20 MINUTES

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Distinguishing between transformed and non-

transformed cells:• Typically involves incorporating an antibiotic resistance gene in the plasmid and then plating the cells on a medium containing that antibiotic.

• Only the transformed cells are resistant, so only they can grow on the medium.

Grow on Selective Medium

Transformed BacteriaWith Fluorescence

Transformed BacteriaGrowth w/o Fluorescence

No TransformationNO Growth

No TransformationGrowth

pGlo – GfpGreen fluorescent protein

Fluorescent• In the laboratory,

fluorescence is easily achieved by exposing the protein to long range UV light or “ black" light.

• The fluorophore absorbs light in the UV-B region (395 nm.. plus a smaller absorbance peak at 470 nm)

• It emits light (fluoresces) at 509 nm, which is in the green part of the visible spectrum

Gfp and Land Mines• Neal Stewart at the

University of North Carolina is developing plants that can detect land mines

• Plants could be ideal biosensors for land mines as seeds would be spread widely and evenly in a suspect field

• The gene that can announce the presence of land mines is gfp

• The gene will be expressed in the presence of a land mine

Green Fluorescent Protein and Plants

GFP and mice

Glo fish• Fluorescent zebra fish

were specially bred to help detect environmental pollutants. By adding a natural fluorescence gene to the fish, scientists are able to quickly and easily determine when our waterways are contaminated

pGlo• Transformation of E. coli

with the pGlo plasmid• Ori• Gene for Gfp• The plasmid contains the

genes for the Arabinose promoter

• The plasmid contains the genes for ampicillin resistance

• If the bacterium uptakes the plasmid it should glow in response to long range uv light

Arabinose operon• araO1 is an operator site. AraC binds to

this site and represses its own transcription. In the presence of arabinose, however, this site helps to activate expression of the PBAD promoter.

Trasnformation movie

P Glo transformation

• Pick one colony from the starter plate.• Use the sterile loops• Swirl the loop in ice cold CaCl2 ( experimental)• Place in ice for 10 minutes ( Your tubes will be incubating when you enter

the room). I have found that a longer incubation period here increases the yield of transformants

• While the tubes are incubating label your plates• LB AMP these plates eliminate bacteria that do not have gene for antibiotic

resistance to ampicillin• LB/AMP? Ara- These plates contain Arabinose and Ampicillin• These are called the selection plates. The Arabinose will induce the gene to

be turned on• LB- Luria Broth Agar – all bacteria should grow on this agar

Heat Shock

• Leave cells in transformation solution on ice for ten minutes

• Transfer to water bath at 42oC for 90 seconds

• Return cells to ice

Recovery and Plating• Incubate bacteria in Luria Broth for

10 minutes before plating in Petri Dish

• Plate your bacteria+ pGlo – LB AMP and LB/Amp/Ara- pGlo – LB and LB/AMP