Biotechnology and Gene Technologies Flashcards
What are the advantages of asexual reproduction?
- It is quick, allowing organisms to reproduce rapidly and so take advantage of resources in the environment.
- It can also be completed if sexual reproduction fails or is not possible.
- All offspring have the genetic information to enable them to survive in their environment.
What are the disadvantages of asexual reproduction?
- It does not produce any genetic variety, so genetic parental weakness will be in all the offspring.
- Not able to adapt to environmental changes.
What is the definition of vegetative propagation?
Production of structures in an organism that can grow into new individual organisms. These offspring contain the same genetic information as the parent and so are clones of the parents.
What is an example of a plant that can asexually reproduce?
The english elm tree
How are the english elm tree able to adapt to reproduce asexually?
When there are damages to the parent plant (e.g disease or burning)
New growths in the form of root suckers to basal sprouts, appear within 2 months of the damage of the main trunk. They grow from meristem tissue close to the ground.
What are the advantages of elm vegetative propagation?
- Root suckers help the elm spread because they can grow all around the original trunk. (clonal patch)
What are the disadvantages of elm vegetative propagation?
When dutch elm disease hits the trees, the new trees got to about 10cm in diameter and became infected and died. Because they are clones they do not have any resistance to the disease.
- No genetic variation so natural selection cannot occur.
How are farmers able to artificially propagate valuable plants?
- Taking cuttings
- Grafting
- Tissue culture
Why is tissue culture better?
Used to generate large stocks of a particularly valuable plant very quickly, with the added advantage that these stocks are known to be disease-free.
What are the steps of micropropagation?
- A small piece of tissue is taken from the plant to be cloned (explant).
- The explant i placed on a nutrient growth medium.
- Divide into a mass of undifferentiated cells called a callus.
- Callus are placed on a growth medium containing plant hormones that encourage shoot growth.
- Transferred to a different one.
- Transferred to a greenhouse before being planted outside.
What are the advantages of plant cloning in agriculture?
- High-quality crops in terms of yield and resistance produced.
- Know what plant is going to be produced.
- Costs are lower
- Quicker
What are the disadvantages of plant cloning in agriculture?
- Equally susceptible to any new pest, disease or environmental change.
What is reproductive cloning?
- Whole individual animal is cloned from another animal.
- The purpose of reproductive cloning is to generate individuals with desirable features.
What is non-reproductive cloning?
- This is when embryonic cells could be used to clone specific tissues and organs.
- The purpose of non-reproductive cloning is to treat damage caused by diseases or accidents.
What is embryo splitting?
- Nucleus comes from an egg fertilised during IVF.
- Embryos produced are clones of the original zygote.
- Mitochondrial DNA will be identical in each clone and identical to the original egg cell.
What is nuclear transfer?
- Nucleus comes from an adult cell.
- Embryo produced are clones of the donor.
- Mitochondrial DNA will not be identical to the donor organism.
What are examples of non-reproductive cloning?
- The regeneration of heart muscle cells following a heart attack.
- The repair of nervous tissue destroyed by diseases such as MS.
- Repairing the spinal cord of those paralysed by an accident that resulted in a broken back or neck.
What are advantages of animal cloning?
- High-value animals, for example cows giving high milk yield, can be cloned in large numbers.
- Rare animals can be cloned to preserve the species.
- Genetically modified animals- for example sheep that produce pharmaceutical chemicals in their milk- can be quickly reproduced.
What are disadvantages of animal cloning?
- High-value animals are not necessarily produced with animal welfare in mind. Some strains of meat-producing chickens have been developed that are unable to walk.
- Excessive genetic uniformity in a species makes it unlikely to be able to cope with, or adapt tom changes in the environment.
- It is unclear that they will be healthy in the long term.
Why are microorganisms often used in biotechnology?
-Grow rapidly = large quantities of product in a short time
- Produce pure products (often more pure than from chemical processes) = less down stream processing
-Not dependant on climate so can grow anywhere
- Easy to genetically engineer = make many products /human products
- Do not require high temperatures = low costs
- Easy to harvest products = low costs
- Can grow on waste materials = low costs & environmental advantages
What are the phases of a standard growth curve?
Lag
Log
Stationary
Death
What happens in the lag phase?
- Organisms are adjusting to the surrounding conditions
- Cells active but not dividing so population fairly constant
- Synthesis of inducible enzymes and factors involved in cell division
What happens in the log phase?
-Population doubles with every generation o High levels of nutrients o Low levels of waste o low levels of competition o few limiting conditions
What happens in the stationary phase?
-Birth rate = death rate so population is stable
o Nutrient levels are dropping
o Waste levels are rising
o Competition is rising
What happens in the death phase?
-Greater number dying than being produced
o Nutrient levels are low
o Waste levels are high
o Competition is high
State the properties of primary metabolites.
Produced as part of normal growth o Essential for life o Produced in line with growth curve o Example: insulin / amino acids / fatty acids / lipids / enzymes. o Produced using a continuous culture o Highest production in the Log phase.
State the properties of secondary metabolites.
Not produced as part of the normal growth
o Not essential for life.
o Only start to be produced in the stationary phase when nutrients are in short supply and competition is high.
o Example: penicillin
o Produced using a batch culture.
o Highest production in stationary phase.
What are facts about batch culture?
Used to produce secondary metabolites
o Everything sterilised and added at the start then left.
o Small quantities of nutrients are added throughout to maintain the stationary phase.
-Not too much = log phase
- Not too little = death/decline phase
What are facts about continuous culture?
Used to produce primary metabolites
o Everything sterilised and added continuously to maintain the log phase.
o Oxygen, p.H and temperature are constantly monitored
Does BC or CC have a better growth rate?
CC because nutrients are added and waste removed continuously, whereas BC nutrients are decreasing and waste is increasing.
Does BC or CC have a better efficiency?
BC- Less as not in operation all the time
CC- More as in operation all the time
Does BC or CC have better yield?
BC- Only 1 batch is lost
CC- Huge volumes of product is lost
Is BC or CC easier to set up?
BC- Easy to set up and maintain
CC- Difficult to setup and maintain
What can you control to manipulate growth conditions in a fermenter?
Limit contamination Monitor pH Monitor temperature Monitor oxygen levels Monitor nutrient levels
What happens when there is too much contamination?
o Kills culture o Spoils product o Produce toxins harmful to health o Competes with inoculants for space, nutrients etc o Decreases the yield of the product
What is asepsis?
o The absence of contamination from unwanted foreign microorganisms or pathogens.
What are aseptic techniques?
o Any technique/manipulation of equipment or materials that are designed to prevent contamination by foreign and unwanted microorganisms o Includes Autoclave – steamed at 121oC for 15 minutes Gloves Disinfectant Fume cupboard Stainless steel tables Flaming equipment Cultures are kept covered Sterilised nutrient mediums
Which four ways can enzymes be immobilised?
Covalent bonding
Entrapment
Adsorption
Membrane separation
What is adsorption?
Attached to an insoluble material such as clay, resin
Held by ionic bonds / hydrophobic associations
Doesn’t affect reaction rate
Can be easily lost by leakage
What is covalent bonding?
Attached using a cross-linking agent using covalent bonds Little leakage of the enzyme
Can have a large loss of function through immobilisation
What is membrane separation?
Trapped behind a partially permeable membrane / microspheres
Allows substrates and products to pass through it but not the enzyme
What is entrapment?
Inside gel beads using alginate / silica Behind a cellulose fibre network Enzyme trapped in natural state Can reduce reaction rate due to diffusion of substrates through immobilising agent Can support enzymes natural state
What are the advantages of large scale immobilised enzymes?
- Does not mix with / does not contaminate / stays separate from, the product; ref to, no / less / easier, downstream processing
- Recoverable / not lost during processing
- Reusable / cost effective = less downstream processing
- Matrix stabilises / protects the enzyme
- So activity not affected by changes in, temperature / pH or run at a high temperature / wider range of pH
- Longer, use / shelf-life
- So suitable for continuous culture / cost effective / greater yield
What are the disadvantages of large scale immobilised enzymes?
o Can reduce the reaction rate due to reduced ability to form enzyme-substrate complexes
o Difficult and costly to immobilise
o If contamination occurs you must destroy all the enzymes
o Can lose enzymes by leakage
What process allows us to make a ‘print’?
Gel electrophoresis
What is the process of electrophoresis?
- DNA samples are treated with restriction enzymes to cut them into fragments.
- Samples placed into wells cut into one end of the gel.
- The gel is immersed in a tank of buffer solution and an electric current is passed through the solution for a fixed period of time.
- Attracted to positive electrode ( shorter move faster)
- Dye is used to stain the DNA molecules.
Outline how the polymerase chain reaction (PCR) can be used to make multiple copies of DNA fragments.
- Artificial DNA replication
- Generates multiple copies of a DNA sample.
- Can only copy pieces of DNA a few 100 bases long.
- Needs a primer (ss DNA 1-20 bases long) to allow DNA polymerase to bind.
What is the first step in PCR?
Mix DNA sample, free DNA nucleotides, DNA primers and DNA Polymerase.
What is the second step in PCR?
Heat components to 95C (1 min)
- breaks the hydrogen bonds between complementary bases on each strand.
- Makes the DNA single stranded.
What is the third step in PCR?
Cool to 55C (45 secs)
- Primers anneal to the 3’end
- Taq polymerase can now bind to primers.
What is the last step in PCR?
Heat to 72C (2 mins)
- Optimum temperature for DNA polymerase.
- A new strand of DNA is synthesised from each template strand.
What processes are used in sequencing the genome of an organism?
PCR and gel electrophoresis
How do you sequence the genome- the entire set of human genes up until inserted into bacteria?
- Genome mapped using micro satellites
- Genome is sheared into smaller fragments.
- Inserted into BACs then into E.coli and cultured.
What happens after the BACs have been inserted into E.coli?
- DNA is then extracted, one BAC at a time.
- Cut into smaller fragments using restriction enzymes. (base sequence worked out)
Outline how gene sequencing allows for genome-wide comparisons between individuals and between species.
- Identifies genes essenital for life
- Highlights evolutionary relationships.
- Model effects of mutations on genes.
- Highlight what genes cause pathogenic effects
- Identification of genetic disorders.
What does recombinant mean?
DNA from 2 or more different sources joined together.
What is genetic engineering?
Involves the extraction of genes from one organisms, or the manufacture of genes, in order to place them in another organism such that receiving organism expresses the gene product.
How many ways can you obtain a gene of interest?
3
What is method 1 of obtaining a gene?
- It can be located using probes
- Cut out of the genome using restriction enzymes.
- Leaving sticky ends
What is method 2 of obtaining a gene?
DNA sequence known- gene can be sequenced.
Sticky ends are added which will be complementary to the ones formed when the vector is cut.
What is method 3 of obtaining a gene?
- Isolate mRNA from the transcribe gene.
- Use the enzyme reverse transcriptase.
- Make single stranded DNA copy of the mRNA strand
- DNA polymerase will make it double stranded.
- Sticky ends are added which will be complementary to the ones formed when vector is cut.
How do you cut a vector?
- Use restriction enzyme to cut the vector.
- Sticky ends form
How do you place the gene into a vector?
- The complementary sticky ends line up.
- Hydrogen bonds form between the to sticky ends.
- DNA ligase seals the sugar-phosphate backbone.
How do you insert the vector into the recipient cell? (6)
- Ca2+ are used with heat shock
- Electroporation
- Microinjection
- Viral transfer
- Liposomes
- Ti Plasmids
Outline how genetic markers in plasmids can be used to identify the bacteria that have taken up a recombinant plasmid.
- Genetic markers are sequences of DNA that are associated with particular traits.
- Insulin production : tetracycline and ampicillin resistance (genetic markers)
Outline the process involved in the genetic engineering of bacteria to produce human insulin. (7)
- Identify the gene for insulin
- Extract the gene
- Revere transcriptase cut plasmid and insulin.
- The gene and plasmid should have complementary sticky ends.
- Insert gene into plasmid (recombinant DNA)
- plasmid uptake by bacteria
- Identify those with insulin and reproduce.
Describe the advantage to microorganisms of the capacity to take up plasmid DNA from the environment.
- Plasmid DNA usually codes for antibiotic resistance.
- Transferring the plasmid between individuals of the same and even different species increased the chances of survival.
Outline how animals can be genetically engineered for xenotransplantation.
By allotransplantation and xenotransplantation.
What is allotransplantation?
- Transplanting cells, tissues or organs between individuals of the same species.
- Can be rejected
What is xenotransplantation?
Transplantation of cells, tissues or organs between animals of different species.
What are the physiological problems with using pigs?
- Different organ size
- Life span shorter
- core body temperate 39C
What are the ethical/medical problems with using pigs?
- animal welfare
- religious beliefs
- disease transfer.
Explain the term gene therapy.
- Therapeutic technique where the functional allele of a particular gene is placed in cells of an individual lacking functioning alleles of that particular gene.
- Can be used to treat recessive conditions but not dominant.
Methods of gene therapy
- Placing a functional allele of the gene.
- Using interference RNA which can bind to mRNA and science the effect of the gene.
- Killing specific cells by adding genes so the cells express proteins which make the cell vulnerable to attack form the immune system.
What is somatic cell gene therapy?
Body cells= specialised
All contain a full chromosome set but most are turned off.
What is gremlin cell gene therapy?
Engineering a gene into a sperm, egg or zygote cell.
All future cells also have trait.
What are the problems with somatic gene therapy?
- Delivery into the recipient cell is tricky.
- Treatment is short lived and needs to be repeated regularly.
- Viruses trigger immune response/ Liposomes are no efficient.
- Modifications are limited to the patient.
What are the pros of germline cell gene therapy?
- Straight forward delivery as techniques are not cell specific
- Modifications are found in future generations
What are the pros of germline cell gene therapy?
- Unethical to engineer embryos
What are the ethical concerns raised by genetic manipulation of microorganisms?
Benefits: - produce human proteins Risks: -containment/escape -mutations=pathogens -antibiotic resistance
What are the ethical concerns raised by genetic manipulation of plants?
Benefits: -golden rice -pest resistance -herbicide resistance Risks: -Less genetic variation -Toxic varieties and allergic reactions -Features passed on to weeds
What are the ethical concerns raised by genetic manipulation of animals?
Benefits: -Increase milk/meat yield -Xenotransplantation -Pharmaceuticals in milk Risks: - Animal welfare -Religious beliefs
What are the ethical concerns raised by genetic manipulation of humans?
Benefits: -Gene therapy Risks: -Unpredictable effects -Germline gene therapy is making decisions for future generations. - Designer babies
