6.1.3

Question 1

What is genetic engineering?

Answer 1

The manipulation of an organism’s genome to achieve a desired outcome

Question 2

What is a transgenic organism?

Answer 2

An organism that carries a gene from another organism

Question 3

What is the first stage of successful genetic modification?

Answer 3

Isolation of the desirable gene

Question 4

What is the most common technique used to isolate the desirable gene?

Answer 4

Use enzymes called restriction endonucleases to cut the required gene from the DNA of an organism. Each type of endonuclease is restricted to breaking the DNA strands at specific base sequences within the molecule

Question 5

In what way do the restriction endonucleases cut the DNA?

Answer 5

Some make a clean, blunt-ended cut in the DNA, but many of them cut the two DNA strands unevenly, leaving one strand longer than the other. These regions with unpaired, exposed bases are called sticky ends. The sticky ends make it much easier to insert the desired gene into the DNA of a different organism. Another technique is the isolation of mRNA for the desired gene and using reverse transcriptase to produce a single strand of complementary DNA

Question 6

What are the most commonly used vectors in genetic engineering?

Answer 6

Bacterial plasmids

Question 7

What is recombinant DNA?

Answer 7

A new combination of DNA from two sources

Question 8

How is the plasmid cut open?

Answer 8

Using the same restriction endonuclease as used to isolate the DNA plasmid

Question 9

What happens after the complementary bases of the two sticky ends are lined up?

Answer 9

DNA ligase forms phosphodiester bonds between the sugar and the phosphate groups on the two strands of DNA, joining them together

Question 10

What marker genes are generally used?

Answer 10

Genes causing a colour change or fluorescence

Question 11

What is transformation?

Answer 11

The transfer of the plasmid with the recombinant DNA into the host cell

Question 12

Give one method of transformation

Answer 12

Culturing the bacterial cells and plasmids in a calcium-rich solution, and increasing the temperature. This causes the bacterial membrane to become permeable, and the plasmids can enter

Question 13

What is electroporation?

Answer 13

Electroporation is another method of transformation where a small electrical current is applied to the bacteria. This makes the membranes very porous so the plasmids move into the cells

Question 14

What is electrofusion?

Answer 14

A way of producing GM cells

Question 15

Describe the process of electrofusion

Answer 15

Tiny electric currents are applied to the membranes of two different cells. This fuses the cell and nuclear membranes of the two different cells together to form a hybrid cell, containing DNA from both. It is used successfully to produce GM plants

Question 16

How is electrofusion used differently in animals?

Answer 16

Animal cells do not fuse as easily and effectively as plant cells. Their membranes have different properties and hybrid animal cells usually don’t survive in the body of a living organism. However, electrofusion is useful in the production of monoclonal antibodies

Question 17

What are examples of engineering in prokaryotes?

Answer 17

Bacteria and other microorganisms have been GMed to produce many different substances that are useful to people. These include hormones, such as insulin and human growth hormone, clotting factors for haemophiliacs, antibiotics, pure vaccines, and many of the enzymes used in industry

Question 18

Describe one method of genetically modifying plants

Answer 18

A desired gene is placed in the Ti plasmid of Agrobacterium tumefaciens along with a marker gene. This is then carried directly into the plant cell DNA. The transgenic plant cells form a callus, which is a mass of GM plant cells, each of which can be grown into a new transgenic plant

Question 19

How can transgenic plants be produced by electrofusion?

Answer 19

The cells produced have chromosomes from both of the original cells and so are polyploid (hybrid). The main stages involve removal of the plant cell wall by cellulases, electrofusion to form a new polyploid cell, the use of plant hormones to stimulate growth of a new cell wall, callus formation, and the production of many cloned transgenic plants

Question 20

What is the genome?

Answer 20

The genome of an organism is all of the genetic material it contains

Question 21

What are the 5 stages of DNA profiling?

Answer 21

1. Extraction of DNA
2. Digesting the sample
3. Separating the DNA fragments
4. Hybridisation
5. Seeing the evidence

Question 22

Describe the process of DNA extraction

Answer 22

The DNA is extracted from a tissue sample using PCR (polymerase chain reaction)

Question 23

Describe the process of digesting the sample

Answer 23

The strands of DNA are cut into small fragments using restriction endonucleases. Two cuts are made, one through each strand. Restriction endonucleases give scientists to cut the DNA strands at defined points in the introns.

Question 24

Describe the process of separating the DNA fragments

Answer 24

Cut fragments of DNA are separated via electrophoresis. The single-stranded DNA fragments are then transferred onto a membrane for Southern blotting

Question 25

Describe the process of hybridisation

Answer 25

Radioactive or fluorescent DNA probes are now added in excess to the DNA fragments on the membrane. They bind to the complementary strands of DNA under particular conditions of pH and temperature.

Question 26

Describe seeing the evidence

Answer 26

The results are observed, in order to view the DNA profile. It is unique to every individual except identical siblings

Question 27

What is PCR?

Answer 27

Polymerase chain reaction is a version of the natural process by which DNA is replicated, allowing scientists to produce a lot of DNA from the tiniest original sample.

Question 28

How is PCR prepared?

Answer 28

The DNA sample to be amplified, an excess of the four nucleotide bases, smaller primer DNA sequences, and DNA polymerase are mixed in a vial placed in a PCR machine.

Question 29

What happens during step 1 of PCR?

Answer 29

The strands are separated, as the temperature inside the PCR machine is increased to 90-95 C for 30 seconds. This denatures the DNA by breaking the hydrogen bonds holding DNA strands together, so they separate.

Question 30

What happens during step 2 of PCR?

Answer 30

The primers are annealed, as the temperature is decreased to 55-60 C and the primers bind to the ends of the DNA strands.

Question 31

What happens during step 3 of PCR?

Answer 31

DNA synthesis. The temperature is increased again to 72-75C for at least 1 min. This is the optimum temp for DNA polymerase to work best. DNA polymerase adds bases to the primer, building up complementary base strands of DNA and so producing double-stranded DNA. Taq polymerase is used.

Question 32

What is a primer?

Answer 32

A short, single stranded, DNA fragment

Question 33

Why is non-coding DNA used in DNA profiling of humans?

Answer 33

In most people, the genome is very similar, so using coding sequences would not provide unique profiles. Non-coding DNA provides variable numbers of short tandem repeats

Question 34

What are short tandem repeats?

Answer 34

Short tandem repeats occur when a short sequence of DNA is repeated many time sin a row.

Question 35

Describe how nucleic acid fragments are separated by electrophoresis

Answer 35

DNA fragments are put into wells in gel strips which also contain a buffer solution. When an electric current is passed through the electrophoresis plate, the DNA fragments in the wells at the cathode end move through the gel towards the positive anode at the other end. The rate of movement depends on the mass of the DNA fragments. Smaller ones travel faster

Question 36

What is Southern blotting?

Answer 36

A technique where the DNA fragments, once denatured, have the strands transferred to a nylon membrane, which is placed over the gel. The membrane is covered with several sheets of dry adsorbent paper, drawing the alkaline solution containing the DNA through the membrane by capillary action,

Question 37

What are the basic principles of DNA sequencing?

Answer 37

The DNA for sequencing is mixed with a primer, DNA polymerase, an excess of normal nucleotides, and terminator bases. The mixture is placed in a PCR machine. Each time a terminator base is incorporated instead of a normal nucleotide, the synthesis of DNA is terminated as no more bases can be added. This results in many DNA fragments of different lengths. The DNA fragments are separated by electrophoresis. The order of baes in the capillary tubes shows the sequence of the new, complementary strand of DNA made.

Question 38

What is bioinformatics?

Answer 38

The development of the software and computing tools needed to organise and analyse raw biological data

Question 39

What is computational biology?

Answer 39

The study of biology using computational techniques

Question 40

What has genome sequencing allowed for?

Answer 40

- Analysis of the human genome - Analysis of pathogenic genomes - Identifying species - Searching for evolutionary relationships

Question 41

What is somatic cell gene therapy?

Answer 41

The replacement of a mutant allele with a healthy allele in the affected somatic cells. Viral vectors are often used

Question 42

What are the drawbacks of somatic cell gene therapy?

Answer 42

It is only a temporary solution for the treated individual, as the healthy allele will be passed on every time a cell divided by mitosis but somatic cells have a limited life. They are replaced from stem cells, which will have the faulty allele

Question 43

What is germ line gene therapy?

Answer 43

The insertion of a healthy allele into an embryo immediately after fertilisation during IVF. The individual would be born healthy and would pass the healthy gene to their offspring

Question 44

What are the concerns surrounding germ line gene therapy?

Answer 44

The potential impact on an individual of an intervention on the germ cells is unknown. Also, the human rights of the unborn individual are arguably violated as it is done without consent. Additionally, this technology could be used to choose desirable or cosmetic characteristics of their offspring.