3.5 genetic modification and biotechnology

Describe the role of restriction enzymes in nature and in biotechnology applications.

Restriction enzymes are proteins that cut DNA at specific sequences. In nature, they protect bacteria from viral DNA. In biotech, they are used to cut and paste DNA for gene cloning and genetic engineering.

Contrast sticky and blunt ends restriction endonuclease.

Explain the function and purpose of DNA electrophoresis.

Electrophoresis is used to separate molecules according to their size and/or charge. The result is a series of "bands" that each contain molecules of a particular size.

The band pattern can be used to identify individuals for:
-forensic analysis
-paternity testing
-determining evolutionary relationships
-testing for alleles associated with disease

Describe how and why DNA fragments separate during electrophoresis.

DNA is loaded into the well of an agarose gel. Because DNA is negatively charged (due to the phosphate groups) it will be pulled through the gel by an electric field.

DNA fragments move towards the positive electrodes, with smaller fragments moving further (depending on mass and charge)

Outline the functions of the buffer, marker and loading dye in DNA electrophoresis.

Buffer: Maintains pH and provides ions for conductivity.

Marker: Indicates DNA fragment sizes for comparison.

Loading Dye: Helps visualise DNA fragments during electrophoresis.

State the function of the PCR.

The Polymerase Chain Reaction is used to make many copies ("amplify") of a specific region of DNA, making millions of copies of a particular DNA sequence.

Describe the selectivity of the PCR.

A specific section of DNA can be copied using PCR.

By using primers that are specific to a certain sequence of nucleotides, only the targeted region will be copied.

Outline the process of DNA profiling.

1. A sample of DNA/blood/saliva/semen is obtained

2. A comparison or reference sample of DNA is also obtained

3. PCR is used to produce more copies of the DNA

4. DNA is cut into fragments by restriction enzymes

5. DNA fragments are separated by size via electrophoresis to form a series of bands

6. bands are compared between the different samples

7. If the banding pattern is the same, then the DNA is from the same source. Children will share bands with either 1 or both parents.

Outline how the universality of the genetic code allows for gene transfer between species. 

The universal genetic code is a common language for almost all organisms to translate nucleotide sequences of DNA and RNA to amino acid sequences of proteins. Because organisms all use the same code, a gene can be removed from one species, inserted into another and the recipient species will transcribe and translate the gene to create a functional protein.

Contrast sexual and asexual reproduction.

Asexual reproduction generates offspring that are genetically identical to a single parent. In sexual reproduction, two parents contribute genetic information to produce genetically unique offspring.

Define “clone”.

A clone is an organism or cell, or group of organisms or cells, produced asexually from one ancestor, to which they are genetically identical.

Define “cloning”.

Cloning means to make an identical copy of [a DNA sequence, cell, tissue or organism].

Outline an example of natural cloning in plants.

Some plants produce clones naturally by asexual reproduction. For example:

Spider plants grow new plants, called plantlets, on their stems

Potato plants produce tubers, which can grow new roots and shoots

Strawberry plants grow stems called runners, which have plantlets on them

Outline an example of natural cloning in animals.

Natural clones, identical twins, occur in humans and other mammals. These twins are produced when a zygote splits, creating two or more embryos that carry almost identical DNA. Identical twins have nearly the same genetic makeup as each other, but they are genetically different from either parent.

Animals such as hydra create clones through a process of budding. A bud develops as an outgrowth due to repeated cell division at one specific site. These buds develop into tiny individuals and, when fully mature, detach from the parent body and become new independent individuals.

Describe the process of cloning via somatic cell nuclear transfer.

Somatic cell nuclear transfer is a technique for cloning. The nucleus is removed from a healthy egg. This egg becomes the host for a nucleus that is transplanted from another cell, such as a skin cell. The resulting embryo can be used to generate embryonic stem cells with a genetic match to the nucleus donor (therapeutic cloning), or can be implanted into a surrogate mother to create a cloned individual, such as Dolly the sheep (reproductive cloning).

Outline the production of Dolly the sheep using somatic cell nuclear transfer.​

List example sources of DNA that can be used in DNA profiling.

DNA can be obtained from:
-blood
-semen
-saliva
-tissue samples

Describe a technique for genetic modification including plasmids, restriction enzymes, reverse transcriptase and ligase.

Gene transfer takes a gene from one organism and inserts it into another.

1. Plasmid removed from bacteria;

2. Plasmid cleaved/cut open by restriction enzymes;

3. The same a restriction enzyme then cuts out a target gene of interest (i.e. the insulin gene) from a chromosome;

4. The plasmid and target DNA are mixed; the sticky ends of the plasmid and target gene of interest will bond at their complementary base sequences;

5. DNA ligase is used to bond the target gene into the plasmid, forming a recombinant plasmid.

6. The recombinant plasmid is inserted into a host cell bacterium.

7. The bacteria with the recombinant plasmid multiply and produce a tiny amount of the protein product (i.e. insulin) from the gene of interest.

Outline why plasmids with genes coding for antibiotic resistance are chosen as vectors in gene transfer between species.

Some, but not all bacteria will accept a recombinant plasmid into their cell. How do we know which bacteria have the taken in the recombinant plasmid and which ones didn't? By using a plasmid with a gene for antibiotic resistance, the bacteria can be grown on a growth medium that included an antibiotic. Only bacteria containing the recombinant plasmid can survive; the rest will die.