4.4 Genetic Engineering and Biotechnology

Polymerase Chain Reaction (PCR):

• technique used to replicate small sections of DNA
• limited samples of DNA can be replicated and studied

Steps of PCR: Automatically alternates between 3 temperatures, repeats and exponentially replicates DNA

1. denature DNA sample into 2 strands. (95 degrees Celsius)
2. annealing, primers bind to regions of DNA in preparation for replication (53 degrees Celsius)
3. primer extended using a heat tolerant DNA polymerase to assemble strands of nucleotides based on the DNA template (73degrees Celsius)

Gel Electrophoresis: 

Can you determine who the criminal is?

• Charged molecules separated by electric field
• Different samples of DNA are put into wells cast in gel. The gel is put into a conducting fluid and then an electric charge is applied
• DNA sample will move throught the gel depending on the charge and size of the molecules. Large molecules will move less distance than small molecules 
• The gel is stained so that the molecules can be seen
• Bands will have appeared according to the sample of DNA and the movement of the molecules. Samples with bands of similar distances means the molecules are approximately the same size and a conclusion can be made that they are from the same source 

Gel electrophoresis can be used in forensices, molecular biology, genetics, microbiology and biochemistry. The gel can be analyzed under a UV light and a gel imaging device. The device compares the intensity and distance of the band compared to standards set on the gel. 

Profiling individuals can be done by using the DNA. Since DNA is unique, each person will have a unique DNA profile. Satellite DNA is made up of highly repetitive regions of DNA to create a long stretch of DNA (What is repeating? The nucleotides, A, T, G, C  or a small sequence of these base pairs might be repeating for an extended length). . The short repetitive regions are called short tandem repeats (STR). Individuals will have DNA with a different number of repeats within the satellite DNA to give specific DNA profiles that can be compared using gel electrophoresis. Gel electrophoresis will determine the density of these repeating regions

The Human Genome Project:

The Human Genome Project was an international project with the goal of sequencing the 3 billion base pairs in the human genome. 

The project was able to find:

• Mapping: discovered the location, number and basic sequence of the human genes. (around 25,000 genes)
• Screening: the sequences has made finding genetic diseases and carriers easier using specific gene probes
• Medicine: New proteins and their functions were discovered that can help improve medicine. The HGP can find the points of the DNA that cause or contribute to disease so that it can be treated, cured or prevented in the future. 
• Ancestry: further insight into the history and evolution of humans

Gene Transfer:

The genes discovered through the HGP are universal. The amino acid sequences which the DNA will code for is the same in all organisms with the rare exception of a few genes. Therefore the DNA of one organism can be transferred and theoretically used by another organism to produce the same protein. Gene transfer could act as therapy for diseased persons by inserting the genes that will produce proteins or hormones to fight the disease. 

Gene transfer basically requires the creation of re-combinant DNA. This is DNA from two or more sources. A gene is inserted into a host cell, a vector, so that when the vector is replicated so will its DNA. After some time, there will be many copies of the foreign DNA. Vectors are typically a bacterial plasmid which are cirlces of DNA not found in the bacterial chromosome.   

1. Isolating Genes for transfer: Restriction enzyme cuts the section of DNA from the genome. This might leave some exposed bases that are called sticky ends
   The plasmid (circle of DNA) is taken from the bacterial cell.
   The plasmid and the isolated DNA can be amplified using PCR
2. Vector preparation for transfer: Same restriction enzyme can be used to cut the plasmid that leaves sticky ends complementary to the isolated gene.
3. Gene Transfer: ligase will glue the sticky ends together to from a continuous strand of DNA
4. The recombinant plasmid is reinserted into the host cell. The host bacteria cell expresses the gene. The cell and its DNA replicates and can be isolated and used. 

Genetically modified organisms:

GMO's are considered organisms that have had their DNA altered through the use of recombinant DNA. Many foods and animals have been genetically modified in hopes of improving the species. 

Golden rice is an example of genetic engineering that introduces genes from daffodil plants and bacteria into rice. This gene is rich is beta-carotene which is the precursor to vitamin A.

Fish tomato is another example of a GMO. It combines a gene from fish into a tomato so that it is resistant to freezing. 

Sheep have been modified to produce more wool by engineering the gene for the enzyme that produces cysteine. Cysteine is found in the keratin protein of wool that keeps wool stronger. 

Possible Benefits of GMOs:

• Characteristics are introduced into organisms that would never have the possibility of obtaining such a trait
• Production of food increases due to the modification
• Can reduce the need for chemical pesticides and therefore the cost of farming. Cheaper costs
• Resistance to different environments can allow certain organisms to grow in new places

Possible Harms of GMOs:

• Unknown effects cause by this new string of DNA, such as allergies or mutations
• Improved plant may compete against or overtake native species 
• Cross pollination where the gene is introduced to weeds making it more difficult to kill the weed

Transgenic Animals:

Genes from a human can be injected into the egg of an animal. The egg can then be implanted into a surrogate mother so that it's offspring, the transgenic animal, will produce the desired trait. 

Benefits of transgenic animals:

• Production of various nutrients, drug and transplant organs that can be used medically
• Introduction of favorable/improved traits and therefore more genetically desirable animals. Traits such as resistance to disease or environmental factors, production of industrial products such as silk or as a food source

Cloning: Clones are organisms that have identical genes or cells that have been taken from a single parent cell

How its done: 

• Cells from one adult is fused with an empty unferttilized egg from a second adult. The embryo created from this fusion can be inserted into the womb of a third surrogate adult. The surrogate mother will give birth to the clone of the donor adult. 
• Broken down: A female animal is stimulated to produce eggs. The nucleus is removed from the egg
• The nucleus of a somatic cell from another animal is taken and fused with the empty egg cell
• Electricity is used to stimulate egg division. As it begins to divide, it is planted into the uterus of a surrogate mother
• The embryo will be a clone to the mother that provided the somatic cell.