Genetic Technology Flashcards
(128 cards)
1
Q
What is the aim of genetic engineering?
A
To remove a gene (or genes) from one organisms and transfer it to another so that the gene is expressed in its new host
2
Q
What is recombinant DNA?
A
- It is DNA made by joining pieces from two or more different sources
- The DNA that has been altered by this process and which now contains length of nucleotides from two different organisms is called recombinant DNA (rDNA)
- The organism which now expresses the new gene or genes is known as a transgenic organism or a genetically modified organism (GMO)
3
Q
Why is genetic engineering important?
A
- It provides a way of overcoming barriers to gene transfer between species
- Unlike selective breeding where whole sets of genes are involved, genetic engineering often results in the transfer of a single gene
4
Q
What is the overview of gene transfer?
A
- GMO produced
1. The gene that is required is identified and it may be cut from a chromosome, made from mRNA by reverse transcription or synthesised from nucleotides
2. Multiple copies of the gene are made using PCR
3. The gene is inserted into a vector, which delivers the gene to the cells of the organisms e.g. plasmids, viruses and liposomes
4. The vector takes the gene into the cells
5. The cells that have the new gene are identified and cloned
5
Q
What is needed for gene transfer?
A
- Enzymes, such as restriction endonucleases, ligase and reverse transcriptase
- Vectors, including plasmids and viruses
- Genes coding for easily identifiable substances that can be used as markers
6
Q
What are restriction endonucleases?
A
- A class of enzymes from bacteria which recognise and break down the DNA of invading viruses known as bacteriophages (phages for short)
- Bacteria make enzymes that cut the phage DNA into smaller pieces
7
Q
What is the function of restriction enzymes?
A
- These enzymes cut the sugar-phosphate backbone of DNA at specific places within the molecule
- This is why they are known as endonculeases (‘endo’ means within)
8
Q
What is the role of restriction enzymes in bacteria?
A
To restrict a viral infection
9
Q
How does each restriction enzyme work?
A
- Each restriction enzyme binds to a specific target site on DNA and cuts at that site
- These target sites, or restitution sites, are specific sequences of bases
- E.G. The restriction enzyme, BamHI always cut DNA where there is a GGAATCC sequence on one strand and its complementary sequence CCTAGG on the other
10
Q
How is DNA protected from such an attack by restriction enzymes?
A
- By Chemical markers
- Or by not having the target sites
11
Q
How do restriction enzymes cut?
A
- Straight across the sugar phosphate backbone to give blunt needs
- In staggered fashion to give sticky ends
12
Q
What are sticky ends?
A
- Short lengths of unpaired bases
- They are known as sticky ends because they can easily form hydrogen bonds with complementary sequences of bases on other pieces of DNA cut with the same restriction enzyme
- When long pieces of DNA are cut with a restriction enzyme, there will be a mixture of different lengths
13
Q
How do you find the specific piece of DNA required?
A
- Separate the lengths of DNA using gel electrophoresis and using gene probes
- Multiple copies of the required piece of DNA can be made using PCR
14
Q
Why is gene tech important?
A
- Now that many proteins have been sequenced, it is possible to use the genetic code to synthesise DNA artificially from nucleotides rather than cutting it out of chromosomal DNA or making it by reverse transcription
- Genes, and even complete genomes, can be made directly from DNA nucleotides without the need for template DNA
- Scientist can do this by choosing codons for the amino acid sequence that they need
- The sequence of nucleotides is held in a computer that directs the synthesis fo short fragments of DNA
- These fragments are then joined together to make a longer sequence of nucleotides that can be inserted into plasmids for use in genetic engineering
- This method is used to generate novel genes that are used in e.g. the synthesis of vaccines
15
Q
How do you get a new gene into a recipient cell?
A
A vector has to be used
16
Q
What are plasmids?
A
- A vector used
- Small, circular pieces of double stranded DNA
- Plasmids occur naturally in bacteria and often contain evens for antibiotic resistance
- They can be exchanged between bacteria. even between different species of bacteria
- If a genetic engineer inserts a piece of DNA into a plasmid, then the plasmid can be used to take the DNA into a bacterial cell
17
Q
How to you obtain the plasmids from bacteria?
A
- Bacteria containing them are treated with enzymes to break down their cell walls
- The ‘naked’ bacteria are centrifuged, so that the relatively large bacterial chromosomes are separated from the much smaller plasmids
18
Q
How is the circular DNA of the plasmid cut open? What is this also used for?
A
- A restriction enzyme
- The same enzyme as the one side to cut out the gene should be used so that the sticky ends are complementary
19
Q
What happens if a restriction enzyme is used that gives blunt ends?
A
-Sticky ends need to be attached to both the gene and plasmid DNA
20
Q
What happens to the open plasmids and length of DNA?
A
- They are mixed together
- Some of the plasmid sticky ends pair up with the sticky ends of the new gene (hydrogen bonding)
21
Q
What is DNA ligase used for?
A
- To link together the sugar phosphate backbones of the DNA molecule and the plasmid, producing a closed circle of double stranded DNA, containing the new gene
- This is now recombinant DNA
22
Q
How can bacterial plasmids be modified to produce good vectors or made artificially?
A
- The pUC group of plasmids have:
1. A low molecular mass, so they are readily taken up by bacteria
2. An origin of replication so they can be copied
3. Several single target sites for different restriction enzymes in a short length of DNA called a polylinker
4. One or more marker genes, allowing identification of cells that have taken up the plasmid
23
Q
What other vectors are there to plasmids?
A
- Viruses
- Liposomes, which are tiny spheres of lipid containing the DNA
24
Q
How do you get the plasmids into the bacteria / get the bacteria to take up the plasmids?
A
- The bacteria are placed in a solution with a high concentration fo calcium ions
- Then cooled and given heat shock treatment to increase the chances of the plasmids passing through the cell surface membrane
- A small proportion of the bacteria, perhaps 1% take up plasmids with the gene, and are said to be transformed
- The remaining either take up plasmids that have closed without incorporating a gene or do not take up any plasmids at all
25
How do you identify the bacteria with recombinant DNA?
1. Spread bacteria on agar plates each containing an antibiotic
2. So if, for example, the insulin gene has been inserted into the plasmid at a point in the gene for tetracycline resistance in pBR322, then any bacteria which had taken up plasmid with the recombinant DNA would not be able to grown on agar contains tetracycline
26
Why is this important to identify the bacteria with recombinant DNA?
Important to identify which bacteria have been successfully transformed so that they can be used to make the gene product
27
What happens after the plasmids have been taken up?
- DNA polymerase in bacteria copies the plasmids
- The bacteria then divide by binary fission so that each daughter cell has several copies of the plasmid
- The bacteria transcribe the new gene and may translate it to give the required gene product e.g. insulin
28
What is a form of diabetes caused by?
- One form of diabetes mellitus is caused by the inability of the pancreas to produce insulin
- Before insulin from GM bacteria became available, people with his from of diabetes were treated with insulin started from the pancreases of pigs or cattle
- Then came the idea of inserting the gene for human insulin
29
What were the problems to producing insulin?
- Locating and isolating the gene coding for human insulin from all the rest of the DNA in a human cell
- Instead of cutting out the gene from the DNA in the relevant chromosome, researchers extracted mRNA
30
What is the initial step in insulin production?
- Extract mRNA for insulin the pancreatic beta cells, which are the only cells that express the insulin gene
- These cells contain large quantities of mRNA for insulin as they are its only source in the body
31
Why do they used insulin mRNA over insulin DNA?
- There are several copies of the mRNA without the non-coding sort of the DNA available which have already been transcribed
- DNA is a long molecule and hard to find wanted section
32
What happens to the extracted mRNA? What is reverse transcriptase and DNA polymerase?
1. The mRNA is then insulated with the enzyme reverse transcriptase (this comes from a group fo viruses called retroviruses)
2. The enzyme reverses transcription using mRNA as a template to make a single stranded DNA (cDNA)
3. These single stranded DNA molecules using DNA polymerase to assemble nucleotides to make the complementary strand
- The genetic engineers now had insulin genes that they could insert into plasmids to transform the bacterium
33
What is the advantage of using GM insulin?
- There is a reliable supply available to meet the increasing demand
- Supplies are not dependent on factors such as availability through the meat trade
- Can also change nucleotide sequence of the insulin gene to give molecules with different aa sequences to give e.g. act faster or slower
34
What are is GFP as a genetic marker?
- GFP (green fluorescent protein) that uses enzymes that produce fluorescent substances
- Enzymes obtained from jelly fish make protein GFP, that fluoresces bright green in UV light
35
What is the concern of using antibiotic resistance genes as markers?
-Antibiotic genes may spread to other bacteria, potentially producing strains of pathogenic bacteria leading to diseases that may be untreatable
36
How does GFP genetic markers work?
- The gene for the enzyme is inserted into he plasmids
- To identify bacteria that have taken up the plasmid, need to shine UV light onto them
- Glow green = GM ones and the same marker gene can be used in a range of organisms
37
What is Beta-glucurondiase (GUS) work as a genetic marker?
- Originates from E.Coli
- Any transformed cell that contains this enzyme, when incubated with seem specific colourless or non-flurescnet substrates, can transform them into coloured or florescent product
- Especially useful in detecting the activity of inserted genes in plants
38
What does a promoter do?
1. Controls expression of a gene
| - The region of DNA to which RNA polymerase bind and starts transcription
39
Why are promoters important in genetic technology?
- If you want the gene that we are going to insert into a bacterium to be expressed, then you also need to insert the appropriate promoter
- When bacteria were first transformed to produce insulin, the insulin gene was inserted next to the beta galactosidase gene so they shared a promoter
- The promoter switched on the gene when the bacterium needed to metabolise lactose
- So if the bacteria were gown in the medium containing lactose but no glucose, they synthesised both beta galactodisae and human insulin
40
What are the functions of promoter?
- The promoter allows RNA polymerase to bind to DNA and it also ensure that it recognises which of the two DNA strands is the template tsrand
- Within the sequence of nucleotide in the promoter region is the transcription start point (the first nucleotide of the gene to transcribed)
- In this way the promoter can be said to control the expression of a gene and can ensure a high level of gene expression
- In Eukaryotas, various proteins known as transcription factors are also required to bind to the promoter region or to RNA polymerase before transcription can begin
41
What is gel electrophoresis?
- A technique that is used to separate different molecules
| - Used in analysis of proteins and DNA
42
What is involved in gel electrophoresis? What does movement depend on?
- Place a mixture of molecules into wells cut into agarose gel and applying an electric field
- The movement of charged molecules within the gel in response to the electric field depends on net (overall) charge, size, composition of the gel
43
How does net overall charge affect ge?
- Negatively charged molecules move towards the anode
- Positively charged molecules move towards the cathode
- Highly charged molecules move faster than those with less overall charge
44
How does size affect ge?
-Smaller molecules move through the gel faster than larger molecules
45
How does composition of gel affect ge?
- Common gels are polyacrylamide for proteins and agarose for DNA
- The size of the 'pores' within the gel determines the speed with which proteins and fragments of DNA move
46
How does electrophoresis of proteins occur?
1. The charge on proteins is dependent on the ionisation of the R groups on the amino acid residues
2. Some R groups are positively charges (-NH3+), (-COO-)
3. Whether these R groups are charged or not depends on the pH
4. When proteins are separated by electrophoresis, the procedure is carried out at a constant pH using a buffer solution
5. Usually proteins have a net negative charge
47
How is ge used in polypeptides?
1. Ge has been used to separate the polypeptides produced by different alleles of many genes
2. E.G Allozymes are variant forms of enzymes produced by different alleles of the same gene
48
How can ge be used to diagnose sickle cell anaemia?
- There are many variant of haemoglobin
1. Adult haemoglobin is composed of four polypeptides (2 alpha-globins and 2 beta globins)
2. In sickle cell anaemia a variant of beta globin has an amino acid with a non-polar R group instead of one with an R group that is charged
3. These two variants can be separated by electrophoresis because they have different net charges
4. This means that haemoglobin molecules in people who have sickle cell anaemia have a slightly lower negative charge than normal haemoglobin
5. Therefore the molecules do not move as far through the gel as molecules of normal haemoglobin
- The test to find out whether someone carries the sickle cell allele makes use of this difference
49
What is electrophoresis of DNA?
1. DNA fragments carry a small charge thanks tot he negatively charged phosphate groups
2. In DNA electrophoresis, these fragments move through the gel towards the anode
3. The smaller the fragments, the faster they move
50
How is electrophoresis of DNA used?
Genetic profiling (fingerprinting) in forensic science
51
What are VNTRs?
1. A region of DNA that is known to vary between different people
2. These regions often contain variable numbers of repeated DNA sequences are are known as variable number tandem repeats
3. Only identical twins share all their VNTR sequences
52
How is genetic profile carried out?
- A region of DNA that is known to vary between different apple is chosen (VNTRs)
- DNA can be extracted e.g. saliva, spot of bold
- Usually quantity of DNA is increased using PCR, which makes many copies of the DNA that has been found
- The DNA is then chopped into pieces using restriction enzymes known to cleave it close to the VNTR regions
- Now the DNA is ready for electrophoresis
- When the current is turned off the gel contains DNA fragments that have ended up in different places
- These fragments are not visible straight away
53
How do you make the fragments visible in electrophoresis of DNA?
1. The fragments are carefully transferred onto absorbent paper, which is placed on top of the gel
2. The paper is then heated just enough to make the two strands in each DNA molecule separate from one another
3. Short sequences of single stranded DNA called probes are added
4. The probes also contains a radioactive phosphorus isotope so when the paper is placed on an. X-ray film, the radiation emitted by the probes (which are sick to the DNA fragments), make the film go dark
5. So we end up with a pattern of dark stripes on the film matching the position that the DNA fragments reached on the arose gel
- Alternatively the probes may be labelled with a fluorescent stain that shows up when UV light is shone onto them
54
What are probes?
- Short sequences of single stranded DNA
| - They have base sequences complementary to the VTNR regions
55
What is PCR and what is it used for?
1. Polymerase chain reaction
2. Method for rapid production of a very large number of copies of a particular fragment of DNA
- Virtually unlimited quantities of a length of DNA can be produced from the smallest quantity of DNA (even one molecule)
56
What are the steps of PCR?
1. DNA is extracted
2. DNA is heated briefly to denature the DNA, which separates the double helix, and leaves the bases exposed
3. Primer DNA is added after cooling and complementary base pairing occurs
4. DNA polymerase uses free nucleotides to synthesise complementary strands
5. The gene has been copied and forms part of two DNA molecules
6. Heating denatures the DNA, which starts a new cycle
57
What does DNA polymerase do?
Used to build new strands of DNA against the exposed ones
58
What is a primer?
-A short length of DNA, often about 20 base pairs long and that has a base sequence complementary to the start of the part of the DNA strand that is to be copied
-The primer attaches to the start of the DNA strand
and then the DNA polymerase continues to add nucleotides all along the rest of the DNA strands
59
What temperature and why are the three stages of copying?
1. Denaturing the double stranded DNA molecules to make sing-stranded ones requires a high temperature, around 95 degrees
2. Attaching the primers to the ends of the single-stranded DNA molecules (known as annealing) requires a temperature of about 65 degrees
3. Building up complete new DNA strands using DNA polymerase (known as elongation) requires a temperature of around 72 degrees
- The DNA polymerases used for this process come from microorganisms that have evolved to live in hot environments
60
What are microarrays?
1. Microarrays have proved a valuable tool to identify the genes present in an organism's genome and to find out which genes are expressed within cells
2. They have allowed researchers to study very large numbers of genes in a short period of time, increasing the information available
61
What is a microarray?
- A microarray is based no a small piece of glass or plastic usually 2cm^2
- Short lengths of single stranded DNA are attached to this support in a regular two dimensional pattern, with 10,000 or more different positions per cm2
- Each individual position has multiple copies of the same DNA probe
- It is possible to search databases to find DNA probes for a huge range of genes
- Having selected the gene probes required, an automated process applies those probes to the positions on the microarray
62
What happens when microarrays are used to analyse genomic DNA?
1. The probes are from known locations across the chromosomes of the organisms involved and are 500 or more base pairs in length
2. A single microarray can even hold probes from the entire human genome
63
How are microarrays used to compare the genes present in two different species?
1. DNA is collected from each species and cut up into fragments and denatured to give lengths of single stranded DNA
2. The DNA is labelled with fluorescent tags so that e.g. DNA from one species may be labelled with green tags and DNA from the other species may be labelled with red tags
3. The labelled DNA tags are mixed together and allowed to hybridise with the probes on the microarray
4. Any DNA that does not bind to probes on the microarray is washed off
5. The microarray is then inspected using UV light which causes the tags to fluoresce
64
What happens when specific tags fluoresce red or green?
- Where this happens, we know that hybridisation has taken place because the DNA fragments are complementary to the probes
- Green and red fluorescent spots indicate where DNA from one species only has hybridised with the probes
65
What happens when tags fluoresce yellow?
- Where DNA from both species hybridise with probe, a yellow colour is seen
- Yellow spots indicate that the two species have DNA with exactly the same base sequence
- This suggests that they have the same genes
66
What happens when tags do not fluoresce?
-If there is no colour for a particular position on the microarray it means that no DNA was hybridised with the probe and that a particular gene is not present in either species
67
What happens after the tags fluoresce?
1. The microarray is then scanned so that the data can be read by a computer
2. Data stored by the computer indicate which genes are present in both species, which genes are only found in one of the the species and which genes are not present in either species
68
What can microarrays be used for?
1. Microarrays also make it possible to detect which genes are being expressed at any specific time in each cells in the body
- EG the genes that are expressed in a cancer cell are different from those active in non-cancerous cells
69
What is another use of microarrays?
1. Microarrays are used to compare which genes are active by identifying the genes that are being transcribed into mRNA
- The mRNA from the two types of cell is collected and reverse transcriptase are used to convert mRNA to cDNA
2. As the quantity of mRNA in a cell at one time is quite small, the quantity of cDNA may need to be increased by PCR
3. The cDNA is labelled with fluorescent tags, denatured to give single stranded DNA and allowed to hybridise with probes on the microarray
4. Spots on the microarray that fluroesece indicate the genes that were being transcribed in the cell
- The intensity of light emitted by each spot indicates the level of activity of each gene
- A high intensity indicates that many mRNA molecules were present in the sample, while a low intensity indicates that there were very few
5. The results therefore not only show which genes are active but also their level of activity and this information is changing the way in which cancers are treated
70
What is bioinformatics?
The is the collection, processing and analysis of biological information and data using computer software
71
Describe bioinformatics
1. Gene sequencing is the determination of the order of base paring
2. This technique has generated hug quantities of data
3. There is also a vast quantity of data about the primary structure, shapes and functions of proteins
4. Bioinformatics combines biological data with computer technology and statistics. It builds up databases and allow links to be made between them
5. The databases hold gene sequences of complete genome, amino acid sequences and protein structures. This allows comparisons to be made with other known genomes. Close similarities indicate recent common ancestry
6. Information can allow the development of drugs such as vaccines against infectious diseases
72
What are the products made by gene technology?
1. Insulin
2. Human growth hormone
3. Thyroid stimulating hormone
4. Factor VIII, a blood clotting protein
73
How can these proteins be produced?
These proteins could be produced using bacteria, yeast and cultures of mammalian cells
74
What are the advantages of using such cells to produce these proteins?
1. These cells have simple nutritional requirements
2. Large volumes of product are produced
3. Production facilities do not require much space
75
What are the practical and ethical problems?
- Few practical and ethical problems because proteins are not extracted from animals or by collecting blood
- The disadvantage is that bacteria do not modify their proteins in the same way that eukaryotes do
76
How do you use eukaryotes for genetic engineering in hamster cells?
1. Genetically modified hamster cells are used to produce factor VIII
2. The gene is inserted into hamster kidneys and ovaries cells and then are cultured in fermenter
3. This avoids the use of donated blood that carried risks of infection
77
How do you use eukaryotes for genetic engineering in transgenic animals?
1. Sheep and goats have genetically modified to produce human proteins in their milk
- E.G Human antithrombin is produced by goats which stops blood clotting
- E.G. Human alpha - antitrypsin is produced by sheep which is used to treat emphysema patients
78
What is cabbage moth caterpillar?
- SCID (servere combined immune-deficency) cannot produce enzyme adenosine deaminase
- This enzyme catalyses the breakdown of deoxyadenosine
- Without this enzymes. build up of toxic metabolites will accumulate and damages the immune system
- This enzyme can be made by inside a genetically modified larva of cabbage moth caterpillar
79
What is somatic cell gene therapy?
Inserting gene into a body cell
80
What is germ (reproductive) cell gene therapy?
Inserting gene into a gamete
- e.g. oocytes could be harvested and then correct allele of CFTR gene could be injected omits am eggs and this egg fertilised by a sperm to produce a zygote
- All cells of the child are produced fringe the GE zygote and therefore will all carry the gene that has been instructed
- And the Childs's gametes will also contain the allele and pass on to their children so the allele is in the 'germ line' spread from generation to generation
- Currently illegal in humans
81
What can gene be genetically engineered to do?
1. Produce vaccines and proteins e.g. albumin
2. Be resistant to herbicides
3. Be resistant to insect pests
4. Contain Vitamin A
82
What is oil rape seed?
1. This has been engineered to be herbicide resistant o the plant is not affected when herbicide is sprayed
2. Herbicide kills any weeds that would compete with the crop for space light water or ions
83
What is oils need rape used as a source for?
- Lubricants
- Human and animal feeds
- Biodiesel fuels
84
What is agrobacterium tumefaciens?
1. Type of soil bacteria often used in genetic engineering
2. Causes gall disease in plants
3. Gains entry to the plants through a wound and stimulates the production of a tumour called a gall in the stem
85
What is the tumour due to?
- The presence of a plasmid called Ti plasmid found inside the bacteria
- Ti plasmid replicates and causes the host plant to release hormones, which stimulate the production of the cells forming the tumour
86
Describe Ti Plasmid
1. Ti plasmid has been isolated and used as a vector to carry genes into a aplenty tissue to create transgenic plants
87
How do you make genetically engineered oilseed rape?
1. Insert desirable gene into a Ti Plasmid
2. Then insert the Ti plasmid into Agrobacterium timefaciens
3. Allow the bacteria containing the Ti palms to infect rapeseed cells
4. Grow the transformed rapeseed cells using tissue culture into whole rapeseed plants
5. Every cell in the plant contained the desirable gene
88
What is the effect of genetically modified plants improve efficiency of uptake of mineral salts?
Reducing fertiliser inputs, so less eutrophication
89
What is the effect of genetically modified plants improve ability to withstand drought or high salt?
Cultivate crop in land where soil/climate is unsuitable
90
What is the effect of genetically modified plants improve resistance to herbicides?
Increase crop yield
91
What is the effect of genetically modified plants improving resistance to disease?
Reduce pesticide input, crop losses reduced
92
What is the effect of improving frost resistance?
Growing and harvest season extended
93
What is genetic screening?
- The analysis of a person's DNA to check for the presence of a particular allele
- This can be done in a duets, in a foetus or embryo in the uterus or in a newly formed embryo produced by in vitro fertilisation
94
What is PGD? What does it involve?
- Pre-implantation genetic diagnosis
1. Mixing the father's sperms with the mother's eggs (coccyges) in a dish (IVF)
2. At the eight cell stage, one of the cells from the tiny embryo was removed
3. The DNA in the cell was analysed and used to predict whether or not the embryo would have a genetic diseases for which both parents were carriers
4. An embryo that was not carrying the allele that would causes the disease was chosen fro implantation, and embryos that did have this allele were discarded
95
What has PGD been used for?
1. Used to avoid pregnancies in which the baby would have had Duchenne muscular dystrophy, thalassaemia, haemophilia, Huntington's disease and others
96
What happened in 2004 with PGD?
It was first used in the UK to produce a baby that was a tissue match with an elder sibling, with a view to using cells from he umbilical cord as a transplant into the sick child
97
What ethical thing happened in 2004 with PGD?
- UK law allows na embryo to be chosen that did not have the allele for a genetic disease and also one that did have a tissue type that would allow a successful transplant into a sick elder brother or sister
- BUT it did not allow the addition of an eel to an egg, sperm or zygote
98
What are some ethical concerns are genetic screening?
1. A fetes can now be screened for a genetic disease while int he uterus using amniocentesis or chorionic villus sampling and the parents may then decide to have the pregnancy terminated if the embryo is found to have a genetic diseases
- Choose termination even if not desired sex, or the 'defect' is relatively minor
2. PGD for sex preselection is considered un ethical
99
What is amniocentesis used for?
- Used to contain a sample of amniotic fluid at 15-16 weeks of pregnancy
- Look for chromosomal mutations
100
What is ultrasound scanning used for?
- To visualise the foetus and to locate the position of the placenta, foetus and umbilical cord
- A suitable point for the insertion of the hypodermic syringe needle is chosen and this is marked on the abdominal skin surface
- Generally this position is away from the foetus, umbilical cord and placenta
101
What is chronic villus sampling used for?
1. Can be carried out between 10 and 13 weeks of pregnancy, so it allows parents to get an easier warning of any genetic abnormalities int he foetus than is possible with amniocentesis
2. Small part of placenta called chorion is removed by a needle and the needles is narrow and the procedure is monitored by ultrasound scanning
102
What is therapeutic abortion?
Terminating pregnancies for a medical reason, rather than for any other
103
What is gene therapy?
Inserting normal alleles of these genes into cells
104
Why is gene therapy harder than initially thought?
-Problem lie in getting normal alleles of the genes into a person's cells and then making them work priorly when they get there
105
What are the most common vectors used in gene therapy?
- The most common vectors that are used to carry the normal allele into host cells are:
1. Viruses (often retroviruses or lentiviruses)
2. Small spheres of phospholipid called liposomes
3. Occasionally naked DNA is used
106
What is SCID?
- Severe combined immunodeficiency
| - Inability to make an enzyme, adenosine deaminase (ADA) which is vital for the functioning of the immune system
107
How has the work with vectors helped in increasing successful gene therapies?
1. The eyesight of young men with a form of hereditary blindness, Leber congenial amaurosis in which retinal cells die off gradually from an early stage as been improves
2. The normal allele of beta globin gene has been successfully inserted into blood stem cells to correct the disorder beta-thalaseeaemia
3. Six people with haemophilia B (in which factor (XI is missing) have at least seen their symptoms reduced
4. Five children were successfully treated for SCID in 2013
108
What is cystic fibrosis?
-A genetic disorder in which abnormally thick mucus is produced in the lungs and other parts of the body
109
What are the effects of cystic fibrosis?
1. A person with CF is very prone to bacterial infections in the lung because it is difficult for the mucus to be removed, allowing bacteria to breed in it
- Need daily therapy to help cough up mucus
2. The pancreatic duct may become blocked and people with cystic fibrosis often take pancreatic enzymes by mouth to help with digestion
3. Around 90% of men with CF are sterile as the thick secretion block ducts in the reproductive system
110
What is the cause of cystic fibrosis?
1. Recessive allele of gene that codes for a transporter protein called CFTR
- This protein sits in the cell surface membranes of cells in the alveoli and other places and allows chloride ions to pass out of the cells
- Faulty version of this protein may be coded for and so does not act properly as a chloride ion transporter
2. CFTR gene found on chromosome 7 and consists of about 250,000 bases
- Someone with one faulty allele is able to make enough CFTR protein to remain healthy
111
How does CF affect you?
1. Normally cells lining the airways and in the lungs pump out chloride ions through the channel in the cell surface membrane formed by CFTR
2. This results in relatively high conc of chloride ions outside the cells and reduced the water potential below that of the cytoplasm cells
3. So water moves out of the cells by osmosis, down eh water potential gradient
4. It mixes with the mucus there making it thin enough for easy removal by the sweeping movements of cilia
5. However someone with CF much less water moves out of the cells, so the mucus on their surfaces stays thick nd sticky
- The cilia, or even coughing can't remove it all
112
What are some options for GT of CF?
- If the normal dominant allele could be inserted into cells in the lungs, the correct CFTR should be made
1. Normal allele inserted into liposomes and sprayed as an aerosol into the Moses
2. Using viruses which were harmless to carry the allele into the passages of the gas exchange system, but some side effects as a result of infection by virus
113
What does herbicide do?
- Fields sprayed with herbicide after the crop has germinate, kill any weeds that would otherwise compete with the crop for space, light water or ions
- This increases the yield of the crop if it is herbicide resistant
114
What does glyphosphate do?
1. Inhibits an enzyme involved in the synthesis of three amino acids and it is absorbed by a plants leave and is transported to the growing tips
2. The amino kids re need for producing essential proteins so the plant dies
3. Various micro-organisms have versions of the enzyme that are involved in the synthesis of these amino acids and the gene that was transferred into crop plants came from a strain of bacterium, Agrobacterium
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What are the most likely detrimental effects of the environment of growing a herbicide resistant crop?
1. The genetically modified plant will become an agricultural weed
2. Pollen will transfer the gene to wild relatives producing hybrid offspring that are invasive weeds
3. Herbicide resistant weeds will evolve because so much of the same herbicide is used
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What are examples of insect-resistant crops?
1. Maize is protected abasing the corn borer, which eats the leaves of the plants and then burrows into eps talk, eating its way upwards until he plant cannot support the ear
2. Cotton is protected against pests such as the boll weevil
3. Insect resistant tobacco, against tobacco bud worm
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What are the most likely detrimental effects not he environment of growing an insect-resistant crop?
1. The evolution of resistance by the insect pests
2. A damaging effect on other species of insects
3. The transfer of added gene to other species of plant
- However less pesticide is sued, reducing the risk of spray carrying to and affecting non-target species of insects in other areas
- Remember that only insects that actually eat the crop are affected
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What is Bt toxin?
1. A gene for a toxin, Bt toxin which is lethal to insects that eat it but harmless to other animals has been taken from a bacterium Bacillus thuringiesis
2. Different strand of this bacterium produce different toxins that can be used against different insect species
3. Crop plants that contain the Bt toxin gene from this bacterium produce their own insecticides
4. However, insect populations can evolve resistance to toxins
5. Large numbers of crop plants containing the genes for Bt toxin may accelerate the evolution of resistance to it
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What are examples of Bt toxin?
1. Many populations of corn borers in the USA are now resistant to Bt toxin
2. The pollen of Bt maize (corn) expresses the gene and has been found to disperse at least 60m wide
120
What are the social conditions with Vitamin A?
1. Where people are poor and rice from the major part of they diet deficiency of Vitamin A is a common and serious proven
2. Vitamin A can cause blindness
3. lack of Vitamin A can cause an immune deficiency syndrome this is a significant cause of mortality in some parts oft he work, especially in children
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What is vitamin A?
- Fat-soluble, vitamin found in oily fish and animal products such as eggs, milk, cheese and liver
- It is also made in our bodies from carotene, the orange carotenoid pigment found inc carrots
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Where are pro-vitamin A carotenoids?
1. Present in the aleurone layer of rice grains, but not in the endosperm, the energy storage tissue in the seed that human eat
2 The aleurone layer is removed from rice when it is polished to produce white rice, but brown rice still contains it
3. The aleurone layer goes rancid if the rice is stored for any length of time which is why white rice is produced and usually eaten instead
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Where did the genes for carotene prosecution come from?
1. Daffodils and a. common soil bacterium, Pantoea ananatis and inserted into rice
2. Further research showed that submitting the gene from the daffodil with one from maize gave even higher quantities of carotene, and the single transformation with these gene is the basis of all current golden rice
- Called golden rice as lots of the orange pigment carotene
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How is golden rice produced?
1. Genes for the production of carotene were extracted fro maize and the bacterium Pantonoea ananatis
2. These genes, together with promoters, were inserted into plasmids
3. The plasmids were inserted into bacteria called agrobacterium tumefaciens
4. These bacteria naturally infect plants and so could introduce the GM modified plasmid into rice cells. They were mixed with rice embryos in Petri dishes, some of which were infected by the bacteria carrying the carotene genes
5. The rice embryos now contains the carotene genes, were grown into adult plants. They produced seeds contains carotene in their endosperm
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What is an example of GM animals?
- GM animals for food production are much rarer than crop plants
- E.G GM Atlantic salmon
- Growth hormone regulating gene from a Pacific Chinnook salmon and a promoter form another species of fish were injected into fertilised egg of Atlantic salmon
- As a result salmon grow all year not just spring and summer as growth hormone is produced throughout the year
- Therefore fish reach market size in 18 months rather than three years
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What are concerns of genetically modified crops?
1. The modified crop plants may become agricultural weeds or invade natural habitats
2. The introduced gene(s) may be transferred by pollen to wild relatives whose hybrid offspring may become more invasive
3. The introduced gene(s) may be transferred by pollen to unmodified plants growing on a farm with organic certification
4. The modified plants may be a direct hazard to humans domestic animals or other beneficial animals, by being toxic or producing allergies
5. The herbicide can now be used on the crop will leave toxic residues in the crop
6. GM seeds are expensive, as is herbicide and their cost may remove any advantage of growing a resistant crop
7. Growers mostly need to buy seed each season, keeping costs high, unlike for traditional varieties, were the grower kept seed from one crop to sow for the next
8. In parts of the world where a lot of GM crops are grown, there is a danger of losing traditional varieties with their desirable background genes for particular localities and tier possibly unknown traits that might be useful in a word where the climate is changing. This requires a programme of growing and harvesting traditional varieties and setting up a seed bank to preserve them
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Why use fluorescent markers over antibiotic resistance genes?
1. Resistance gene passed to other bacteria
2. Easier to identify
3. More economical/ time saving
4. Antibiotics no longer effective/development of new antibiotics needed
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Why does a promoter need to be transferred with the desired gene?
1. Promoter initiates transcription
2. Binding of RNA polymerase
3. Otherwise gene has to be inserted near an existing promoter
4. Difficult to do/may disrupt expression of existing gene
5. In eukaryotes precise position of promoter is improtnant
