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Genetic Engineering

Ch.12

Genetic Engineering

• Genetic Engineering - It is now possible todirectly alter the genetic makeup of anorganism.– Cutting DNA– Changing DNA– Removing and reinserting DNA

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Restriction Endonuclease• Restriction Endonuclease – Recognizes and cuts

DNA at a particular sequence.– EcoR1, BamI, HaeIII

Restriction Endonuclease

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Restriction Endonuclease• Sticky Ends

– Unpaired DNA bases that result from a cut by a restrictionendonuclease.

– Can be attached to another molecule of DNA cut with the samerestriction endonuclease.

Sticky Ends

Restriction Endonuclease

Sticky Ends

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Restriction Endonuclease

Bacteria• Bacteria

– Prokaryotes - lacks a nucleus– Simplest forms of life– Very small (nanometers)– Reproduce asexually– Most abundant and diverse

organism in the world.– Some are helpful

• Photosynthetic bacteria, bacteriain your large intestine, bacteriaon your skin, bacteria thatdecompose dead organisms….

– Some are harmful• Food poisoning, colds,

infections…

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Bacteria Structure• No membrane bound organelles• Cell wall - maintains the cell’s shape• Capsule - helps the cell attach to surfaces.• Flagellum - some bacteria have one or more.• Pili - helps the cell stick to surfaces• DNA- one molecule of circular DNA

Bacteria Structure• Plasmids – Small circular pieces of DNA found in

bacteria in addition to their chromosomes.– Can be removed from bacteria and cut up using

restriction enzymes.– A DNA sequence can be inserted into a plasmid.– Plasmids can be easily reinserted back into bacteria.

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Bacteria Structure• Plasmids carry genes for…

– resistance to antibiotics– resistance to heavy metals– sensitivity to mutagens– sensitivity or resistance to bacteriophages– production of restriction enzymes– production of amino acids– production of toxins

Cloning Genes By Using Bacteria• Recombinant DNA – Combining DNA

from two different sources.• Transgenic organism - An organism

which contains foreign DNA

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Cloning Genes By Using Bacteria• Transformation-inserting a plasmid which contains a

foreign gene into bacteria– Remove a plasmid from a bacterium– Use a restriction endonuclease to remove a gene from an

organism - Create sticky ends!– Use the same restriction endonuclease to cut open the plasmid-

Create sticky ends!– Insert the gene into the plasmid– Place the plasmid back into the bacterium

Cloning Genes By Using Bacteria• Reasons to do this

– Clone the gene - Many copies of the plasmid are made bythe bacteria as the bacteria reproduce

– Express a protein such as insulin, human growth hormone,interferon.

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Uses For Transgenic Organisms• Transgenic Bacteria

– Researchers may need a particular protein inlarge quantities for research.

– A pharmaceutical may wish to produce largequantities of growth hormone or insulin.

– Bacteria which digests toxins and pollutantssuch as oil and sewage.

Uses For Transgenic Organisms• Transgenic Plants – Genes can be inserted

in to the plant DNA by a bacterium whichinfects plants.– Natural insecticides such as BT– Plants which produce their own fertilizer– Fruits and vegetables which don’t spoil as

quickly.• Transgenic Animals – DNA is inserted

into the nucleus of a fertilized egg cell.– Animals for research.

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Cloning an Entire Organism• Clone – A newborn individual is a genetic

copy of the adult.– Produce large numbers of organisms which

have favorable characteristics.– Save an endangered species

• Remove the nucleus from an egg cell replace the nucleus with a nucleus from anadult organism Place the egg back intothe foster mother.

Cloning an Entire Organism

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Gel Electrophoresis• Used to determine the size of DNA fragments• DNA samples are run through a porous gel called

agarose.• The DNA is pulled through the agarose by running

an electric current through the agarose gel.• DNA has a negative charge• DNA molecules migrate toward the anode which

has a positive charge• Large fragments of DNA move slowly through the

agarose while small DNA fragments move quickly.• A molecular weight marker is used to estimate the

size of the DNA fragments

Gel Electrophoresis

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Gel Electrophoresis

Gel Electrophoresis

• Uses for gel electrophoresis– Identify similarities and differences among

different kinds of organisms.– DNA fingerprinting.– Separate DNA by size.– Determine the sequence of an organism’s DNA

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Polymerase Chain Reaction (PCR)

• PCR = Polymerase chain reaction• A lab technique which produces millions of

copies of DNA.• Can be used for many applications

including– DNA fingerprinting– Diagnosis of diseases

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Gene Therapy• Gene therapy - using

DNA in order to treat adisease.

• A functional genereplaces a mutated gene.

• Gene therapy treatmentsare becoming more andmore successful asresearch progresses.