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what is pencillium chrysogenum
antibiotics
what is fusarium venenatum
single cell protein
what is lactobaciullus bulgaricus
yogurt
what is rennin enzyme
cheese
what is saccharomyces cerevisai
bread and beer
what is escherichia coli
insulin
what is biotechnology
making use of living organisms (or parts of) in industrial processes
when was biotechnology officially named
1919
why has there been recent advancements in biotechnology
because of genetic technologies
bioremediation
the use of either naturally occurring or deliberately introduced microorganisms to consume and break down environmental pollutants, in order to clean a polluted site
requires: microbe which digest the contaminants as the conditions suit the microbe
used to clean soil contamination and underground water
usually carried out in-situ but may sometimes be ex-situ depending on need
soya
soya beans are fermented to produce soy sauce
uses yeast or Aspergillus
what are the arguments for biotech using microbes
avoids alternative chemical processes- which often require higher temperature and pressure
microbes are easy to culture and often digest waste
rapid reproduction increase number of useful microbes
can be run in lab worldwide
pure product which is easy to harvest
which other organisms can be used and how
such as sheep, goats and cows which can be used to produce useful proteins
the proteins are often produced in the milk for easy harvesting
cows have also been used to synthesis human antibodies, which is isolated from their blood
what is fermentation
anaerobic respiration by microbes
what is coagulation
the thickening of milk due to denaturation of proteins used in yoghurt and cheese
what is rennet
chemical containing the enzyme rennin- which causes coagulation of milk proteins (casein)
what is (kappa) casein
it keeps casein (milk protein) soluble
what are single celled proteins
a protein source from microbes- usually fungus
what is mycoprotein
fungal protein
e.g. Quorn
what is the process to make yogurt
it is milk that has undergone fermentation
raw material: milk
microorganism: lactobacillus bulgaricus and streptococcus thermophilus
effect: bacteria convert lactose to lactic acid which causes the milk protein to coagulate. also partially digest the milk make is tart
additional microbes may be added as probiotics to aid gastrointestinal
what is the process to make cheese
raw material: milk
microorganism: Lactobacillus then rennet (contains rennin)
effect: initial coagulation from lactic acid then further coagulation of casein due to calcium ions and breakdown of kappa-casein (keeps casein soluble) by rennet. Casein precipitates of due to calcium ions
separation of curd (solid) pressed into moulds and then further treatment (may be inoculated with mould)
what is the process to make bread
raw material: flour, water, salt
microorganism: fungus, yeast- Saccharomyces cerevisiae
effect: anaerobic respiration of yeast produces carbon dioxide which makes dough rise
process required: mixing, proving (warm place for anaerobic respiration to occur) and cooking (evaporates alcohol)
what is the process to make alcohol
raw material: grapes containing fructose and glucose (wine), barley which is germinating (beer or ale)
microorganism: naturally occurring yeast (S. cerevisiae)
effect: anaerobic respiration of the sugar by the yeast to produce ethanol and carbon dioxide
what is the processes to make single cell proteins
raw material: any organic substrate e.g. waste paper or whey with sugar and nitrogen source
microorganism: Fungus: Fusarium venenatum
effect: Fungus grows on organic material, producing protein. the most common type is Quorn
what are the advantages of using microbes
protein based foods can be made many times faster than using animals or plants
can easily change rate to suit needs
very high protein content
no animal fat or chloresterol
can adjust the amino acid content (so that there are more rare amino acids)
no seasonal variation
not much land
what are the disadvantages of using microbes
some people do not want to eat food grown on waste
the proteins need isolating
high amounts of nucleic acid that needs removing
infection as also ideal conditions for pathogenic organisms
palatability- not the normal taste or texture
what is large scale lab production
it usually takes place in a specially designed fermenter
the process may be batch or continuous fermentation
growing conditions are carefully controlled to maximise yield
aseptic conditions must be maintained to prevent contamination
what variables need to be controlled for fermentation and why?
Temperature- to prevent enzymes being denatured or growth being too slow
Nutrients- carbon, nitrogen, minerals and vitamins needs for growth
Oxygen- want aerobic respiration
pH- enzyme action
Product concentration- could affect rate or reaction
Sterilisation- prevent contamination and competition
Mixing- brings substrates to organisms
how does continuous culture work?
continuous culture involves constantly topping up the materials inside the fermented whilst constantly harvesting the product
this is used for primary metabolites and gives a constant rate of production
how does batch culture work?
batch culture involves putting the microorganism under stress so it produces secondary metabolites
the process is carried out and then the fermenter is emptied to allow harvesting and restarted
what is penicillin?
modern strains of Penicillium chrysogenum have been selectively bred to be more productive
it is regarded as a secondary metabolite as it is only produced above a certain population size. this means the continuous method cannot be used
how is penicillin made into the drug?
the fermenter is run for 6-8 days. the culture is then filtered to remove the cells
the antibiotic is precipitated as crystals by the addition of potassium compounds. it is then modified
it is then mixed with inert substances to be administered in tablet form
what is the process to produce genetically modified insulin?
been produced by genetic engineering since 1987
human insulin gene inserted into bacterial plasmid (E.coli)
produced by continuous culture- relatively low cost and vast quantities can be produced to meet the demand
what is a culture?
a population of microbes
what is a colony?
a genetically identical group of bacteria produced from a single bacterium
what is a growth medium?
provide nutrients for microbes to grow
what is agar?
a growth medium containing C and N sources
what is an aseptic technique?
sterile technique with no contamination
what is inoculation?
the introduction of a microbe to a growth medium
what is incubation?
providing ideal conditions for growth (usually warmth)
what is an autoclave?
high temperature and high pressure environment to provide sterile equipment
what are the microbe requirements?
carbon source- yeast extract
nitrogen source- peptones from gelatine breakdown
warmth- from incubation
moisture- water in agar
no competition- from sterile technique
a growth medium will contain these things and may be in liquid broth form or jelly-like agar
what are the basic rules of aseptic technique
you must be as clean as possible (wash hands, lab coat)
work area must be sterile- disinfect, cone of sterility
equipment must be sterile (flame bottles upon opening and closing, keep lid on petri dish, flame glass/ metal equipment)
think about every action in advance
what is the 3 stage process
sterilisation- of nutrient agar and equipment
inoculation- introduction of desired microbe to the growth medium
incubation- creating conditions for microorganisms to multiply
what are the sterilisation steps?
autoclave- high temperatures and high temperatures (121oC, 15 minutes)
used for equipment and medium
this kills all living organisms. once cooled enough the medium is poured into a sterile dish and left
what are the 3 methods of inoculation?
streaking- wire loop transfers drop onto agar
seeding- small drop of liquid culture on surface of agar or in dish before agar
spreading- glass spreader used to spread drop over agar
what are the incubation steps?
leave to settle before taping
do not seal completely- anaerobic
label with date, contents and initials
place in incubator (never above 30 degrees)
store upside down to prevent condensation drips
examine after 24-36 hours
autoclave dishes, never open
what is liquid broth?
must be aseptic
most often used to create a population before transfer to agar
serial dilutions used to allow counting of colonies
how do you measure growth rate?
serial dilutions
the number of microorganisms in a broth can be hard to count. so to investigate growth the population density must be reduced
it is normal to do this by a factor of 10.1 of broth to 9 of distilled water each time
a drop of each solution is used to inoculate agar plate. the one you can count needs multiplying back up
what is lag phase?
small population, low growth rate
adjustment phase: gene activation for metabolism, protein synthesis, cell growth
what is the log phase
this is exponential growth
enzymes, nutrients and space all present. rapid growth and reproduction. every generation doubles in size
what is the stationary phase
when nutrients are becoming limited and metabolic waste products accumulate, growth rates decline until the point that growth rate equals death rate
in this phase there is no increase in the population of live bacteria
generally in this phase bacteria produce endospores, toxins and antibiotics
what is the decline phase?
nutrients are exhausted
lethal concentration of waste products
what are primary metabolites?
collected in the log phase
maintain optimal conditions in continuous fermentation
what are secondary metabolites?
produced at end of stationary phase
closed culture
run to decline phase
what are enzymes in biotech?
both intra and extra cellular enzymes are used in biotechnology
enzymes may be isolated from natural sources or manufactured
in all cases they are reusable after the reaction
immobilisation allows the enzyme to be held outside of the reaction mixture and thus reused easily
what are the advantages of immobilisation?
easier to separate enzyme and products
increases stability and can be manipulated easily
allows continuous production/enzyme used for longer
enzyme can be recovered and reused
enzyme does not contaminate product/no purification required
what are the disadvantages of immobilisation?
immobilisation may alter shape of enzyme
may reduce catalytic ability- not freely mixing with substrate
expensive to set up, time, equipment and materials
contamination is costly
what happens in absorbsion?
where enzymes bind with immobilising support
uses a porous support
clay, resin, glass beads
hydrophobic interactions and ionic links
enzymes can become detached (leakage) easily
active site may be distorted lowering activity
what happens in covalent bonding/cross linking?
covalent bonds link enzyme to an insoluble material
cross-linking agent used
unlikely to become detached
likely to interfere with active site
expensive to produce
what happens in entrapment?
trap enzymes in a gel bead or network of cellulose fibres
does not affect active site
reaction rates can be reduced because active site is less easily available
substrate molecules must also be able to pass through entrapment barrier
what happens in membrane separation?
enzymes are physically separated from substrate mixture by a partially permeable membrane
substrate molecules must be small enough to fit through this membrane
product molecules must also be able to pass back through membrane
active site unaffected
what are the advantages of natural cloning?
conditions that are good for the parents are good for the offspring
rapid as the population can increase rapidly
reproduction can be carried out with one parent and sexual reproduction not possible
what are the disadvantages of natural cloning?
offspring may become overcrowded
no genetic diversity (unless mutation during DNA replication)
little variation
selection not possible
whole population susceptible to changes in the environment
why can plants reproduce by cloning?
because of vegetative propagation, they have many cells that retain the ability to differentiate
what is vegetative propagation?
a method of asexual reproduction in plants where a new plant grows from a fragment of the parent plant
what are examples of vegetative propagation and how do they work?
runners- called a rhizome if underground -horizontal stem growths that can form roots at certain points
suckers- the old branch may die and the new branch replaces it
bulbs- underground system from which a group of fleshy leaves grow
corms- solid rather than fleshy like a bulb
leaves- clones grow on the leaf margins. immature plants drop off and take root
tubers- underground stem. one potato can grow into one or more plants
what is tissue culture?
growing new tissues, organs or plants from certain tissues cut from sample plants
what is micro propagation?
growing large number of plants from meristem tissue taken from a sample plant
when should micropropagation be used?
when the desirable plant doesn’t:
produce many seeds
doesn’t respond well to natural cloning
rare
GM or selectively bred
needs to be pathogen free
what is the micropropagation process?
cells are taken from shoot (called explant)
cells are sterilised before being placed onto nutrient medium
explants placed on sterile growth medium (glucose, phosphates, amino acids, hormones)
forms a callus culture
divided to produce lots of small clumps or undifferentiated cells
transferred to a new agar medium- plantlets grow
plantlets are transferred into compost
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