Chapter 15 Genetic Engineering Cloning and Transgenic Organisms

Applied Genetics Selective Breeding – Desired traits of plants and animals are selected to be passed on to future generations. Hybridization – The crossing of two varieties of a plant or animal to get chosen desired characteristics.

Gene Technology The process of manipulating genes for practical purposes is called genetic engineering Scientists can transfer genes from one organism to another designing new living things to meet specific needs. This process involves building recombinant DNA – DNA made from two or more different organisms

Transgenic organisms Any organism that contains recombinant DNA is a transgenic organism. Transgenic organisms contain foreign DNA in their cells.

Transgenic Bacteria Used to make: –Insulin –Human Growth Hormone –Clotting Factors

Example: Insulin and Bacteria Insulin is a protein hormone that controls sugar metabolism. People with _____________can not produce enough insulin. They need to take doses of insulin. Human gene for insulin is transferred into bacteria. Bacteria can transcribe and translate this gene to make insulin.

How does this happen? 1. Cutting DNA –The DNA from the organism is cut out by restriction enzymes Human DNA is cut to get the gene of interest Bacterial DNA called a plasmid is cut to open it up –Enzyme EcoR1 – A restriction enzyme that cuts DNA at the following sequence of bases…

X X X G A A T T C X X X X X X C T T A A G X X X X X X G A A T T C X X X X X X C T T A A G X X X X X X G X X X C T T A A G X X X A A T T C X X X G X X X EcoR1 cuts at GAATTC CTTAAG ‘sticky’ ends The DNA that has been ‘cut open’ can now receive a new section of DNA from some other organism.

2. Making recombinant DNA –DNA fragment from human is inserted into the plasmid DNA 3. Cloning –Many copies of the gene of interest are made after it has been inserted into a bacterial cell (the host cell) 4. Screening: Host cells that did take up the gene are then separated from the cells that did not. The host cells with the new gene then transcribe and translate the DNA with the inserted gene. They will make insulin!

Transgenic bacteria Genes for making human insulin have been inserted into bacteria.

Transgenic Animals Used to study genes and improve the food supply –Mice with human-like immune systems. –Livestock with extra copies of growth hormone genes. –Sheep and pigs that produce human proteins in their milk.

Example: Spider Goat? -DtkIhttp:// -DtkI

Transgenic Plants Used for food supply and medical supplies –Soy and corn that grow better –Rice with additional Vitamin A –Plants that produce blood proteins –Plants that produce vaccines

Example: GM Crops Genetically modified crops can lead to better, less expensive, more numerous food and less harmful manufacturing processes. Cotton, soybeans and corn have been inserted with a bacterial gene called Bt toxin. –Not harmful to plants and humans and most other animals –Acts as a pesticide, Bt toxin kills insects –Crops don’t have to be sprayed with pesticides, production increased.

Other Transgenic Examples For example the gene that codes for green fluorescent protein (GFP) in jellyfish has been added to tobacco plants, insects and even a pig.

Transgenic Tobacco plant with GFP

Fruit fly with GFP

Transgenic pig with GFP protein

GFP gene inserted in a mouse embryo

PCR Once biologists find a gene (like GFP) they often make copies of it. Why would they do this?

Polymerase Chain Reaction (PCR) Makes copies of a section of DNA This can be used to increase the amount of DNA found at a crime scene Can be used to make copies of a sequence of DNA in order to figure out the order of the nitrogen bases

PCR 1. A piece of DNA is heated and separated into two strands.

2. At each end of the original piece of DNA, a primer is added and it complements a portion of the sequence of DNA

3. DNA polymerase copies the region between the primers. These copies then serve as templates to make more copies. 4. In this way, just a few dozen cycles of replication can produce billions of copies of the DNA between the primers.

Cloning Producing genetically identical organisms from one original cell. We have successfully cloned pigs, cows, mice, sheep, monkeys, cats… The first animal cloned was a sheep named Dolly in 1997.

Dolly the sheep. The first cloned animal. She died of cancer but appeared normal otherwise at her death.

What is cloning? Every single bit of DNA is the same between the two! Natural Example: Identical Twins, Mitosis, binary fission Two Types of cloning “Reproductive” cloning and “Therapeutic” cloning.

Reproductive Cloning performed with the express intent of creating another identical organism. The organism is the exact duplicate of one that already exists or has existed in the past. Cloning of plants, animals, and humans falls into the class of reproductive cloning. uses a technique called somatic cell nuclear transfer (SCNT)

A donor cell is taken from a sheep’s udder. Donor Nucleus These two cells are fused using an electric shock. Fused Cell The fused cell begins dividing normally. Embryo The embryo is placed in the uterus of a foster mother. Foster Mother The embryo develops normally into a lamb—Dolly Cloned Lamb Egg Cell An egg cell is taken from an adult female sheep. The nucleus of the egg cell is removed. Reproductive Cloning

Therapeutic Cloning A cell is removed from a patient requiring medical treatment. The nucleus of the patient’s cell is removed and inserted into an empty donor egg. Cell division is encouraged through the use of special chemicals or an electric current. The egg is allowed to grow into a blastocyst. The resulting stem cells are removed from this blastocyst and used to treat the patient. No rejection since it’s the patient’s own DNA.

A blastocyst – none of these cells have differentiated or specialized into any particular type of cell so they are all stem cells. A cell could be removed and someday used to grow into any type of cell or organ.

Is a clone genetically identical? Why might they look different? Clones would become different as they live in a different environment.

Problems with Reproductive Cloning? Low success rate (Dolly took 276 attempts) Genetic defects such as poor immune systems Massive quantities of eggs required Reduction in adaptability – genetic uniformity Lack of knowledge of how to do it

Is cloning allowed? Different countries have different laws for the different types of cloning. Some countries want a global ban on all types of cloning, while others want a ban on certain types. In South Africa there are currently no laws against cloning.

And lets clone!! Seeing the real thing, enucleating a cell and nuclear transfer.Seeing the real thing, enucleating a cell and nuclear transfer. HHMIwww.youtube.com/watch?v=hepoJgGJtNc HHMI