Enzymes that Manipulate DNA
What is the role of a restriction endonuclease?
Cuts strands of DNA
What is PCR?
- Polymerase chain reaction
- amplifies a sample of DNA
What does gel electrophoresis do?
Separates DNA fragments based on their size
Name the two types of reproductive cloning technologies in animals
Somatic cell nuclear transfer (SCNT)
Embryo splitting
What is a plasmid?
A small, circular loop of DNA found in bacteria
What is genetic engineering?
The alteration of an organism’s genome using genetic recombination technologies
What are stem cells?
Undifferentiated cells that have the ability to differentiate into specialized cells.
Name the virus that typically attacks a bacteria
Bacteriophage
Name the three approaches to bioethics
●Consequences- based
●Duty/ rule- based
●Virtues- based
What is the role of a polymerase?
To 'build' DNA or RNA
What is a primer?
Short, single strand of nucleic acids that acts as a starting point for polymerase enzymes to attach.
Why do DNA fragments move towards the cathode?
DNA is negatively charged due to the presence of the phosphate group. As opposites attract, DNA is going to be attracted to the positive terminal / electrode (cathode)
What is an example of naturally- occurring embryo splitting?
Formation of identical twins
What is the difference between a recombinant plasmid and a non-recombinant plasmid?
A recombinant plasmid contains the gene of interest, whereas a non-recombinant plasmid does not.
Why would scientists want to modify an organism's genome?
So that the organism will have desireable traits that were originally missing
What two features must a cell have to be classified as a stem cell?
- Must be capable of self renewal
- Potency - capable of differentiating into other types of cells
What molecule cuts bacterial DNA and viral DNA?
Cas9 (a type of endonuclease)
Name the five ethical concepts
Integrity
Respect
Beneficience
Non- maleficence
Justice
What is the difference between DNA ligase and RNA ligase?
oDNA Ligase – joins two DNA fragments together.
oRNA Ligase – joins two RNA fragments together.
Why is Taq polymerase used rather than normal DNA polymerase?
Taq polymerase is heat resistant, allowing it to catalyse elongation stage of PCR. DNA polymerase would denature
Explain how you would identify whether one fragment of DNA is smaller or larger than another fragment
The smaller fragments will move further than the larger fragments
Name two examples of cloning in plants
Plant cutting
Micropropogation
Plant grafting
Define bacterial transformation
The process by which a bacteria takes up a recombinant plasmid
What is the difference between a GMO and a TGO?
GMO - an organism that has had its genome modified.
TGO - a type of GMO that has had a gene from a different species inserted into its genome (eg. inserting a bacteria's gene into a moo cow's genome)
What is the advantage of using embryonic stem cells compared to adult stem cells?
Embyronic stem cells can replicate indefinitely and turn into multiple types of cells, whereas adult stem cells cannot replicate indefinitely, and thus can turn into limited cell types
Where does the viral DNA get stored within the bacteria's genome?
In a CRISPR gene
Issues with GMOs can be classified into three main categories. Name and define them.
○Biological – implications relating to nature and its organisms.
○Social – implications relating to the economy, environment, or culture.
○Ethical – implications relating to whether something is morally, and therefore ethically, acceptable.
Where do restriction endonucleases cut DNA?
What bond gets broken in the process?
Recognition sites
Phosphodiester bonds
Outline the process of PCR. Include the name of each stage, the temperature, and an outline of what happens
1 - Denaturation - 90-95oC
DNA is heated to 90-95oC to break the hydrogen bonds between the bases and separate the strands.
Single-stranded DNA forms.
2 - Annealing - 50-55oC
The single- stranded DNA is cooled to allow the primers to bind to complementary sequences on the single- stranded DNA.
3 - Elongation - 72oC
The DNA is heated so that Taq polymerase can function optimally. Taq polymerase binds to the primer, which acts as a starting point, and begins synthesizing a new complementary strand of DNA.
What is a standard ladder? Why is it used?
●A standard ladder contains a number of different DNA fragments of known size.
●We use the standard ladder to identify sizes of unknown bands.
SCNT - which organism will the clone be genetically identical to?
The somatic cell donor
Provide an example of an application of bacterial transformation
- Insulin production
Name and explain two reasons why scientists would modify crops.
oIncreasing crop productivity - increase amount of food made to reduce starvation OR increase nutritional value of food to reduce malnutrition OR increase ability of the crop to grow in adverse conditions (eg. heat) so that more food is produced.
oIncreasing disease-resistance - increases food security by minimising crop destruction
Define:
- Totipotent
- Pluripotent
- Multipotent
Totipotent - Can differentiate into any cell type
Pluripotent - Differentiate into multiple cell types
Multipotent - Can differentiate into limited specialised cell types belonging to a specific tissue/ organ
What does CRISPR stand for?
Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)
What is a bioethical issue?
An ethical dilemma relating to biology that typically involves a decision-making process between two or more options for an action
What is the difference between blunt ends and sticky ends?
Blunt end cuts are made in the middle of a recognition site, making a vertical cut, whereas stick end leave overhanging nucleotides
What is the difference between a forward primer and a reverse primer?
Forward primers:
•Binds to the start codon at the 3’ end of the template strand.
•This causes Taq polymerase to synthesis a new DNA strand in the same direction that RNA polymerase would function.
Reverse primers:
•Binds to the stop codon at the 3’ end of the coding strand.
•This causes Taq polymerase to synthesise a new DNA strand in the reverse direction that RNA polymerase would function.
Name and briefly describe two factors that impact how far DNA fragments move.
●Voltage – the stronger the electric force, the further the DNA travels towards the positive electrode.
●Gel composition – the greater the density and/ or concentration of the adrose, the more difficult it is for larger fragments to move. This means that the DNA will travel a shorter distance.
●Buffer concentration – the greater the concentration of ions in the buffer, the more the electric charge is conducted through the gel. This causes DNA to move further down the lane.
●Time – the longer the electric current is applied, the further the DNA will travel.
Outline the process of Somatic Cell Nuclear Transfer (SCNT)
oEnucleation: removal of the nucleus from the donated egg cell. Enucleated egg cell results.
oExtraction: extract somatic cell’s nucleus.
oInsertion: insert somatic cell’s nucleus into the enucleated egg cell.
oDevelopment: after insertion of the modified egg cell, the cell begins to divide and develop into an embryo, which is then implanted into a surrogate mother.
Name and briefly describe two methods that can increase the likelihood of a bacteria taking up a recombinant plasmid.
What do these methods do to the bacteria to increase the likelihood that the recombinant plasmid is taken up?
- Heat shock - plasmids are placed in a calcium ion solution, and then briefly heated up, before being returned to cold.
- Electroporation - exposing the bacteria to an electric current
- Increases the permeabiltiy of the bacteria's membrane
Bt crops are an example of a GMO. Are Bt Crops an example of a TGO or a cisgenic organism? Explain why by referencing the types of genes inserted into the genomes of Bt crops.
Additionally, explain WHY these crops are produced
- TGO
- Contains a crystal toxin gene from bacteria
- Prevents crop destruction by particular insects
Provide examples of:
- Totipotency
- Pluripotency
- Multipotency
Totipotent - Zygote: can differentiate into placental cells, and cells required to build a fetus
Pluripotent - Embryonic stem cells
Multipotent - Bone marrow: contains blood stem cells. Can differentiate into a variety of blood cells.
Why is a CRISPR gene said to be a palindrome?
Because it can be read the same forwards and backwards
A GMO is an organism whose DNA has been altered using genetic engineering techniques. GMOs are often created so that an organism expresses a particular trait. For example, in agriculture, organisms may be modified to be pest/ pesticide/ disease/ drought- tolerance, produce higher yield, have longer shelf lives, or have a greater nutritional content.
The use of GMOs for human and animal food is a bioethical issue.
Choose two bioethical concepts, and explain how they can be used to inform the debate.
1.Integrity – manufacturers might need to clearly label their products as GMOs, allowing consumers to be better informed.
2.Justice – GMOs might create inequity between larger agricultural companies who have resources and funding to genetically modify their crops, versus a smaller family farm who do not.
3.Beneficence – there are positive health outcomes for people who consumer GM foods that have been nutritionally enhanced.
4.Non- Maleficence – GMOs might cause unintended disruptions to the food web (e.g. insect-resistant crops may alter population levels of different pests).
5.Respect – it is important to promote the right of individuals to freely choose whether or not they use GMOs, and be provided with equation representation of alternatives.