1
Q
What are 3 techniques used to study genes?
A
- The polymerase chain reaction (PCR)
- Gel electrophoresis
- Cutting out DNA fragments using restriction enzymes
2
Q
What is PCR used for?
A
To make multiple copies of (amplify) a DNA fragment
3
Q
What are primers?
A
Short pieces of DNA that are complementary to the bases at the start of the fragment you want
4
Q
What is a palindromic sequence (recognition sequence) of nucleotides in DNA?
A
Where the nucleotides sequences consist of the same order of bases when read backwards on the opposite strand.
5
Q
What are restriction enzymes used for?
A
To get a DNA fragment from an organisms DNA.
6
Q
How do restriction enzymes work?
A
They recognise palindromic sequences (recognition sequences) and cut (digest) the DNA at these places
- Different restriction enzymes cut at different specific recognition sequences, because the shape of the recognition sequence is complementary to the enzymes active site.
- If recognition sequences are present at either side of the DNA fragment you want, you can use restriction enzymes to separate it from the rest of DNA
- The DNA sample is incubated with the specific restriction enzyme, which cuts the DNA fragment out via a hydrolysis reaction.
7
Q
What are sticky ends?
A
- Sometimes the cut from restriction enzymes leaves sticky ends which are small tails of unpaired bases at each end of the fragment.
- Sticky ends can be used to bind (anneal) the DNA fragment to another piece of DNA that has sticky ends with complementary sequences.
8
Q
What method is used to produce DNA profiles?
A
Electrophoresis
9
Q
How is electrophoresis used to produce a DNA profile?
A
- Organisms genome’s consist of repetitive, non-coding base sequences, called tandem repeats, and the number of times these are repeated differs from person to person.
- The number of times a sequence is repeated at different loci in a person’s genome (and so the number of nucleotides) can be analysed using electrophoresis, which creates a DNA profile.
10
Q
Why is DNA profiling used in forensic science?
A
It compares samples of DNA collected in crime scenes to samples of DNA from possible suspects, to link them to crime scenes.
11
Q
How is DNA profiling used in medical diagnosis?
A
- A DNA profile can be a unique pattern of several alleles
- It can be used to analyse the risk of genetic disorders and is useful when the specific mutation isn’t known or where several mutations could have caused the disorder, because it identifies a broader, altered genetic pattern
- It can screen embryos for genetic disorders before they’re implanted into the uterus by IVF. The fault regions of the parents DNA are used to produce DNA profiles, which are compared to the DNA profile of the embryos. If the profiles match, the embryo has inherited a disorder. It can scene for cystic fibrosis, Huntington’s disease, etc.
12
Q
What is genetic engineering?
A
The manipulation of an organism’s DNA
13
Q
What are transformed organisms?
A
Organisms that have had their DNA altered by genetic engineering
14
Q
What is recombinant DNA?
A
DNA formed by joining together DNA from different sources - this is what genetically modified organisms have.
15
Q
What does genetic engineering involve?
A
Extracting a gene from one organism and inserting it into another organism. Genes can also be manufactured instead of extracted. The organism with the inserted gene will then produce the protein coded for by that gene.
16
Q
What is a transgenic organism?
A
An organism that had been genetically engineered to include a gene from a different species.
17
Q
What is the first stage of genetic engineering?
A
The DNA fragment containing the desired gene is obtained, the fragment is isolated using restriction enzymes.
18
Q
What is the second stage of genetic engineering?
A
The DNA fragment (with the gene in) is inserted into a vector, which is used to transfer DNA into a cell.
19
Q
What vectors can be used in genetic engineering?
A
Plasmids or bacteriophages
20
Q
How is the DNA fragment inserted into a vector?
A
- The vector DNA is cut open using the same restriction enzyme that was used to isolate the DNA fragment containing the desired gene.
- This means the sticky ends of the vector are complementary to the sticky ends of the DNA fragment.
- The vector DNA and DNA fragment are mixed together with DNA ligase. DNA ligase joins up the sugar-phosphate backbones of the two. This is ligation.
- The new combination of the bases in the DNA (vector DNA + DNA fragment) is recombinant DNA
21
Q
What is the third stage of genetic engineering?
A
The vector transferring the gene into the bacteria
22
Q
How are plasmid vectors taken in by bacterial cells using electroporation?
A
- A suspension of the bacterial cells is mixed with the plasmid vector and placed in a machine called an electroporator.
- The machine is switched on and an electrical field is created in the mixture, which increases the permeability of the bacterial cells is membranes and allows them to take in the plasmids.
23
Q
How are bacteriophage vectors taken up by bacterial cells?
A
- The bacteriophage will infect the bacterium by injecting its DNA into it. The phage DNA (with the desired gene) then integrates into the bacterial DNA.
24
Q
How is insect-resistance created in plants?
A
Using genetic manipulation to insert a gene into the cells of the plant which makes them resistant to insect plants,
25
Why were soybeans genetically modified to be resistant to some insects?
Soybeans are an important food source across the world, but yields can be greatly reduced by insect pests that feed on the plant. The gene codes for a protein that is toxic to some of the insects that feed on the plant.
26
How are soybeans GM?
- Using restriction enzymes to insert a desire gene from a bacteria into a plasmid. The plasmid is put back into the bacteria. The plant cells are deliberately infected with the transformed bacteria. The desired gene gets inserted into the soybean plant cells DNA.
27
What is a positive of GM plants?
Reduces the amounts of chemical pesticides that farmers use on crops, which can harm the environment.
28
What are negative ethical issues of GM plants?
It may encourage monoculture (where only one type of crop is planted). Monoculture decreases biodiversity and could leave the whole crop vulnerable to disease as all the plants are genetically identical.
29
What is pharming?
When medicinal drugs can be produced using genetically modified organisms, such as animals.
30
How can be pharming carried out?
- DNA fragments that code for production of specific proteins are injected into the animal embryo
- The embryo is implanted into a female, and when the offspring is born it is tested to see if it produces the protein, e.g, in its milk
- If it does produce the protein, selective breeding is used to produce a herd of the animal that produce the protein in their milk
- The protein can be extracted from the milk to produce a drug
31
What are positive ethical issues of pharming?
Drugs can be made in large quantities compared to other methods of production, making them more avaliable to people
32
What are negative ethical issues of pharming?
- Manipukating an animal’s genes could cause harmful side effects for the animal, and using an animal in this way enforces that animals are assets that can be treated however we choose, animals can’t give consent
33
How can genetic engineering be carried out on pathogens to find treatments to disease?
- Bacteria can be genetically modified to produce useful substances to people, like insulin and vaccines
34
What ethical issues does genetic engineering of pathogens raise?
- It has helped to cure diseases, as previously untreatable disease can now be treated, reducing suffering
- However:
- There are worries the scientist researching the pathogen could become infected and cause a mass outbreak of disease
- The genetically modified pathogen could revert back to its original form and cause an outbreak of disease
- In the wrong hands, knowledge of how to genetically engineer dangerous pathogens could be used maliciously to create agents of biowarfare
35
What is patening and technology transfer for genetically modified products?
- Scientists working for different institutions often share their knowledge and skills in the field, so that globally, beneficial genetically modified products can be created at a faster rate. This is technology transfer
- A group of scientists may want to gain legal protection for their genetically modified products, by getting a patent which means by law they can control who uses the product and how.
36
What are the positives of patening?
- This has positives like the owner of the patent getting money from selling the product to develop more methods. It also encourages scientists to compete to be the first to come up with new beneficial genetic engineering ideas
37
What are the negatives of patenting?
There are negatives like farmers in poorer countries not being able to afford patented genetically modified seeds. Some patents also mean they are not legally allowed to plant and grow any of the seeds from the crop without paying again. Some patents mean they can’t plant and grow seeds from the crop without paying again. This is unfair and the big companies that own the patents are benefitting the most without regard to help farmers in poorer countries.
38
What is gene therapy?
Altering alleles to cure genetic disorders
39
How can a genetic disorder caused by two recessive alleles be fixed?
Adding a dominant allele to make up for them
40
How can a genetic disorder caused by a dominant allele be fixed?
- ‘Silence’ the dominant allele e.g. putting DNA in the middle so it doesn’t work anymore
41
How is the new allele inside the cell in gene therapy?
Using vectors, like alternated viruses, plasmids or liposomes
42
What are the two types of gene therapy?
- Somatic therapy
- Germ line therapy
43
What is somatic gene therapy?
- Altering the alleles in body cells, particularly the cells that are most affected by the disorder
- Replacing the mutant allele with a healthy allele in affected somatic (body) cells
44
What are the drawbacks of somatic gene therapy?
- It is only a temporary treatment as somatic cells have a limited life and are replaced by stem cells, which will have the faulty allele
- A treated individual will also still pass the faulty allele onto any children they have
45
What is germ line therapy?
- Altering the alleles in sex cells
- Inserting a healthy allele into the germ cells, or into an embryo immediately after fertilisation
- The individual would be born healthy with the normal allele in place and would pass it into their own offspring
46
What are the positives of germ line cell therapy?
- The individual would be born healthy with the normal allele in place
- The individual would pass the healthy allele onto their own offspring
47
What are positives of gene therapy?
- Prolongs lives of people with genetic disorders
- Gives them a better quality of life
- Carriers might be able to conceive a baby without the disorder with germ line therapy
- Germ line could decrease the number of people that suffer from genetic disorders
48
What are negative ethical issues of gene therapy?
- The treatment could be used in other ways than medical treatment like choosing desirable or cosmetic characteristics in their offspring
- Risk of harm in the patient e.g. overexpression of genes
- Expensive and short lived (somatic)
- Difficult to get into specific body cells
- Body could identify vectors as foreign bodies and start an immune response against them
49
What is the process of sequencing DNA called?
Sanger sequencing
50
What is the method of DNA sequencing?
- A single stranded DNA template, DNA primer, DNA polymerase, free nucleotides and flourescently labelled modified nucleotide (which once it’s added to a DNA strand, no more can be added after) are all added into four separate tubes
- The tubes undergo PCR, which produces many strands of DNA. The strands are different lengths because each one terminates at a different point depending where the modified nucleotide was added.
- The DNA fragments in each tube are separated by electrophoresis and visualised under UV light
- The complementary base sequence can be read from the gel, the smallest nucleotide is at the bottom of the gel. Each band after this represents one more base added. By reading the bands from the bottom of the gel to the top, you can build the DNA sequence one at a time.
51
How can Sanger sequencing be used to sequence whole genomes?
It only works on fragments up to 750 base pairs long, so to sequence the entire genome (all the DNA) of an organism using this method, you need to chop it up into smaller pieces first which are sequences and put back into order to give the sequence of the whole genome.
52
What are the faster, high-throughput DNA sequencing techniques?
- Continued research and improvements in modern technology have led high-throughput sequencing - techniques that can sequence a lot faster than original methods at a fraction of the cost.
- For example, the Sanger sequencing technique has been made high-throughput by new technology allowing hundreds of sequences to be run in parallel.
It has also become automated and is faster - the tube contains all the modified nucleotides, each with a different coloured fluorescent label, and a machine reads the sequence for you. So instead of running a gel and determining the sequence from that, you get a computer read-out.
- There are also newer methods of high-throughput sequencing some of which don't use electrophoresis, for example, high-throughput pyrosequencing. These methods are known as next-generation sequencing and can sequence millions of DNA molecules at the same time — much faster than Sanger sequencing.
4) With newer, faster techniques, scientists can now sequence whole genomes much more quickly.
53
How has sequencing genes led to synthetic biology by showing polypeptide structure?
- Animo acids are coded for by triplets of bases in a gene.
- This means that by sequencing a gene, the sequence of amino acids that a gene codes for and so the primary structure of a polypeptide can be predicted.
- This has allowed us to create biological molecules from scratch and so has led to the development of an area of biology called 'synthetic biology.
- Synthetic biology is a large field that includes:
building biological systems from artificially made molecules (e.g. proteins) to see whether they work in the way we think they do, redesigning biological systems to perform better and include new molecules, designing new biological systems and molecules that don't exist in the natural world, but could be useful to humans, eg. energy products (fuels) and drug products.
54
How can sequences genes and genomes be compared?
- Gene sequences and whole genome sequences can be compared between organisms of different species and betveen organisms of the same species.
- This is a complicated process which is made easier with the use of computers - involves computational biology (using computers to study biology) and bioinformatics (developing and using computer software that can analyse, organise and store biological data).
- There are many reasons why biological research can involve comparison of gene sequences and genomes
55
How has bioinformatics and computational biology contributed to research in genotype-phenotype relationships?
- It's useful to be able to predict an organism's phenotype by analysing its genotype.
- Bioinformatics has allowed the scientists to compare all the data and identify genotype-phenotype correlations — this could help in the treatment of syndromes by using gene sequencing to predict what health problems the person is likely to face.
56
How has bioinformatics and computational biology contributed to research in epidemiological studies?
- Epidemiology is the study of health and disease within a population — it considers the distribution of a disease, its causes and its effects.
- Some gene mutations have been linked to a greater risk of disease.
- Computerised comparisons between the genomes of people that have a disease and those that don't can be used to detect particular mutations that could be responsible for the increased risk of disease.
57
How has bioinformatics and computational biology contributed to research in evolutionary relationships?
- All organisms evolved from shared common ancestors (relatives). Closely related species evolved away from each other more recently and so share more DNA.
- Whole genomes of different species can be sequenced and then analysed using computer software to tell us how closely related different species are. E.g. the genomes of humans and chimpanzees are about 94% similar.
- Comparing the genomes of members of the same species can also tell us about evolutionary relationships. - For example, when different groups of early humans separated and moved to different parts of the world, their genomes changed in slightly different ways. By using computers to compare the genomes of people from different parts of the world, it's possible to build up a picture of early human migration.
