Chapter 22 - Cloning and biotechnology

Question 1

vegetative propagation

Answer 1

➜ plants can reproduce asexually using a cutting (meristem - undifferentiated plant cells that form in the vegetative organs of a plant)

Question 2

Vegetative organs

Answer 2

• root and shoot tips
• axillary buds (where leaves and stem meet)
• Vascular cambium (between xylem and phloem)

Question 3

Process of vegetative propagation

Answer 3

➜ over time a miniature plant (plantlet) forms at the vegetative organs and is attached to parent
➜ these are clones of parent
➜ at maturity plantlet detaches from parent when it is of photosynthesising by itself
➜ new plant = same phenotype so growing and harvesting easier
➜ some plants have runners or horizontal stems that form over soil surface, pointing sufficiently far away from parent so when ready it will grow far away from parent so it doesn’t compete for water or nutrients or is overshadowed by parent

e.g strawberries, peppermint, spider plant

Question 4

adventitious roots

Answer 4

➜ roots forming under nodes of runners
➜ The runner dies when the plantlet is self-sustaining

Question 5

Propagation techniques

Answer 5

➜ dont need seeds
➜ as well as runners, tubers, rhizomes, bulbs, suckers and offsets can be used to propogate
➜ All modes of vegetative propagation contain modified stems that can generate meristematic tissue

Question 6

Potato tubers

Answer 6

➜ swollen modified roots that form eyes on their surface
➜ eyes can sprout new growth = chitting
➜ starch stored in tuber fuels early growth of new plant

Question 7

Rhizomes

Answer 7

➜ Ginger forms rhizomes
➜ it is a modified stem that grows horizontally underground
➜ New growth stems from nodes in the rhizome, forming new stems and adventitious roots
➜ section used in cookery is the rhizome

Question 8

bulbs

Answer 8

➜ onion and garlic
➜ form bulbs that can grow adventitious roots underground and leafy shoots above ground

Question 9

Suckers

Answer 9

➜ growths thatappear from root system of many trees and shrubs which can provide meristematic tissue for vegetative propagation
➜ poplars, cherries and plums

Question 10

Offsets

Answer 10

➜ small virtually complete daughter plants that have been asexually produced on the mother plant
➜ tulips and lillies

Question 11

Practical: Producing Cuttings

Answer 11

➜ Cuttings can grow to full-sized plants much quicker than plants grown from seeds
➜ equipment:
∘ sharp scissors
∘ potting compost
∘ pots to house new plants
∘ hormone rooting powder - contains auxins that promote mitosis and cell differentiation in new root growth
∘ plastic bag to cover cutting
➜ cut below a stem node - small length of stem
➜ dip base of cutting in rooting powder
➜ once new roots have started to grow, it can be planted into soil

Question 12

Cultivars

Answer 12

➜ gardners etc can harness natural process of vegetative repro to artificially clone
➜ like cutting, layering, grafting, division and budding
➜ this creates cultivars - strains of genetically identical plants for foodstuffs or commercial blooms
➜ methods rely on the formation of meristematic tissue from which plant organs can differentiate

Question 13

Cauliflower cloning

Answer 13

➜ many plant cells = totipotent
➜ cauliflower used as it is comprised mostly of actively dividing cells and can withstand being handled
➜ small piece of plant cut = explant
➜ explant then disinfected to prevent fungi
➜ used on endangered species

Question 14

Micropropagation and tissue culture method

Answer 14

➜ wear eye protection for god knows what reason
➜ wipe app surfaces down and soak all apparatus in sterilant (so no fungi contamination)
➜ break off a small floret of cauliflower and use scalpel to cut thin section of floret (5-10mm long) thin section = explant
➜ soak explant in sterilising solution for 15 min and swirl every 5 min = to make sure only cauliflower cells present
➜ take out explant using STERLISISED forceps and add to agar growth medium which has all nutrients needed for plant to grow (agar also sterilised dw)
➜ leave container on sunny window for 3 weeks

RESULTS:
➜ clones a full cauliflower from explant
➜ shows totipotency = explant has capability to produce all diff cel types to make cauli

Question 15

Advantages of plant cloning

Answer 15

➜ plants produced are free of disease
➜ same genotype and phenotype
➜ GM plants are immune to certain diseases
➜ process is rapid so large yield
➜ small plants = easily transported
➜ pants that are difficult to grow via seeds can be cloned
➜ grown in any country = any season
➜ rare and endangered species can be propogated to save them
➜ whole plants can be created from GM cells
➜ use of cultivars prevent risk of F₁ hybrids

Question 16

Disadvantages of plant cloning

Answer 16

➜ expensive and labour intensive process
➜ process is susceptible to microbial contamination
➜ no genetic variation = risk of large scale loss due to disease etc
➜ New plants have to be carefully screened for abnormalities that could lead to the new plants being infected
➜ risk of an unexpected secondary metabolic chem reaction causing stunted growth or evend eath in new explants

Question 17

Asexual reproduction in animals

Answer 17

➜ less common than plants
➜ some reproduce asexually by parthenogenesis - growth and development of embryo clones occur without fertilisation e.g aphids
➜ other naturally occuring cloning = identical twins

Question 18

Identical twins

uno i always wanted a twin 😔

Answer 18

➜ egg fertilised by sperm in a singleton birth
➜ forms zygote
➜ single zygote undergoes a few mitotic divisions to become embryo
➜ identical twins referred as monozygotic
➜ at embryo stage, embryo split into 2
➜ 2 embroyes formed = identical and both develip in utero
➜ result = identical offspring, identical pheno and genotype
➜ non identical twins formed from diff egg and sperm they ARE NOT clones

Question 19

Embryo twinning

Answer 19

➜ produces offspring that are genetically identical to each other and not parents
➜ been routine procedure to boost yield of livestock and promote desired allele since 1980’s
➜ key step is the deliberate division of the emrbyo into 2 half embryos
➜ then inserted into surrogate mum for gestation and birth
➜ births identical twins
➜ some cases - emrbyos are split into single identical cells which can be implanted into separate surrogate mummies
➜ guarantees desirable characteristic in offspring but not how many offspring

Question 20

Reproductive cloning

DOLLY SHEEP IN EDINBURGH, 1996

Somatic Cell Nuclear Transfer (SCNT)

Answer 20

➜ 3 separate animals are required:
∘ animal to be cloned via cell
∘ female to donate egg cell
∘ surrogate mummy
➜ Procedure
∘ animal to be cloned donates a somatic (body) cell
∘ egg cell extracted from donor and enucleated (nucleus is removed by suction and discarded)
∘ nucleus from somatic cell injected into egg cell
∘ hybrid zygote is now treated to encouraged to divide via mitosis
∘ embryo implanted into surrogate mother for gestation and birth

Question 21

Therapeutic cloning

Answer 21

➜ cloned cells are used to replace dead or damaged cells that cause a loss of function in an individual
➜ embryos go through reproductive cloning (dolly sheep process) but the embryos are removed and subdivided
➜ embryo cell = totitpotent so it is artificially differentiated into any type of specialised cell
➜ high potential but little clinical progress cuz UK dum

Question 22

For animal cloning

Answer 22

➜ well accepted and non controversial in livestock farming
➜ maximising agricultural output as desirable characteristics can be cloned
➜ can remove less desirable characteristics from gene pool over time
➜ help preserve endangered species
➜ Provide regenerated organs for patients suffering from degenerative disease - organs would be a direct match so no risk of rejection

Question 23

Against animal cloning

Answer 23

➜ SCNT is hit or miss (100’s of attempts to make dolly bestie)
➜ unknown long term effects of cloning (Subsequent cloning attempts have led to a high number of early deaths and genetic abnormalities in the clones)
➜ some cloned animals tend to grow abonormally large (Large Organ Syndrone - LOS) which can cause breathing and circulatory problems
➜ disrupts normal mechanisms or regulation of gene expression
➜ destroys embryos - ethical?

Question 24

Biotechnology

Answer 24

➜ harnesses the processes in living organisms to useful products like food/meds
➜ carry out useful services liek sewage treatment, composting and bioremediation(use of microorganisms to degrade contaminants of an environmental accident)

Question 25

Microorganisms

Answer 25

➜ most useful group of organisms that carry out biotechnological processes because they
- have simple growth requirements
∘ their food is cheap and readily available
∘ occupy very little space
- reproduce quick
- no non-productive tissues and organs
-can be grown on an industrial scale to perform duties useful to large numbers of the human population

Question 26

Brewing and distilling

Answer 26

➜ brewing uses yeast to respire sugars from barley malt and produce ethanol (primary product) and CO2 (by product)
➜ this is anaerobic and known as fermentation
➜ some alcoholic drinks have a higher alcohol content than fermentation alone can produce, so these drinks are first fermented then distilled to concentrate alc
e.g
∘ whiskey (nasty+it burns) and bourbon are distilled from a barley beer
∘ brandy is distilled from grape wine (yum)
∘ other spirits use generic fermented ethanol distilled through botanicals (berries, herbs etc) to extract flavours

Question 27

Baking bread

Answer 27

➜ wheat/rye mixed with yeast and other stuff to make doUGH
➜ culture of yeast is in fact a mixture of several different naturally occurring species
➜ commercial bakers = control species of yeast used
➜ artisn bakeries use wild yeast cultures, preserved and cultivated regularly
➜ Yeast enzymes begin by hydrolysing the starch in flour to maltose
➜ Maltose is then hydrolysed to produce monosaccharides which can be used for aerobic respiration
➜ O2 runs our so yeast respires anaerobically
➜ aerobic and anaerobic resp produce CO2 (bubbles) throughout dough causing it to rise
➜ baking kills yeast and causes the gas pockets to expand so bread rises more

Question 28

Cheesemaking

Answer 28

➜ pasteurised milk = raw material
➜ bacteria are used to digest lactose producin g lactic acid and it lowers pH of milk
➜ low pH=proteins denature in milk leading do separation of curds (solid) and whey (liquids)
➜ curds are pressed and processed into cheese
➜ mould spores from saprotrophic fungi can be artifically added into blue veined cheeses

Question 29

Yoghurt making

Answer 29

➜ starter culture of Lactobacillus bulgaricus and Streptococcus thermophilus bacteria are introduced to pasteurised milk
➜ bacteria use sugar in milk to respire and produce lactic acid (waste)
➜ lactic acid denatures proteins in milk causing them to coagulate (stick together) producing thick and sour yogurt
➜ flavours added to make it taste yum

Question 30

Penicillin production

Answer 30

➜ species of mould from the Penicillium genus can be cultured in industrial fermenters
➜ known as deep tank fermentation
➜ extractin & purification of product produces large volumes of the drug for therapeutic use
➜ first wonder drug = produced at large scale

∘ penicillin mould added to culture medium with sugars, AA and minerals
∘ stainless steel tank has a water jacket to maintain a constant fermentation temp outside the cultutre medium
∘ bubbles supply O2 and cause mixing of culture and theres a sterile air injection

Question 31

Insulin production

Answer 31

➜ previously diabetes were treated with pig insulin (hard to isolate, expensive and not as effective)
➜ Recombinant DNA technology can incorporate the gene for human insulin into the genome of the bacterium, Escheriscia coli
➜ Recombinant bacteria are grown in batch fermenters, and each bacterial cell expresses insulin
➜ insulin is released into the batch medium and can be purified for medicinal use at a later stage

Question 32

Mycoprotein production

myco means fungus

Answer 32

➜ meat substitute and is low fat and high fibre
➜ microorganism used is Fusarium venenatum, a filamentous fungus
➜ source of glucose is added to the tank
➜ O2 also supplied to allow aerobic resp can occur which yields maximal growth of hyphae
➜ Nitrogen introduced in the form ammonia
➜ product in mycoprotein = fungus, rather than a substance produced by microorganisms

Question 33

Bioremediation

Answer 33

➜ humans contamnate land/water with toxic substances and it can be remediated by removing pollutants and restoring land
➜ e.g oil spills, industrial accidents, acidic damage from mining and cleanup of crime scenes
➜ biore rely on oxidative digestion of pollutants
➜ bioventing (Naturally occurring microorganisms perform aerobic digestion of the contaminants and release non-polluting products) are sometimes enough
➜ biostimulation - relies on naturally occurring microorganisms but adds nutrients that promote microbial diegestion of pollutants
➜ GM has been trialled to create microorganisms capable of biorem

Question 34

Emerging uses of biotechnology

Answer 34

➜ Production of biofuels (to replace the use of fossil fuels)
➜ Production of vaccines and antibodies for the treatment of disease
➜ Production of hardy crop plants that can grow in arid conditions
➜ Counteracting threats from bioterrorism or bio-warfare

Question 35

Advantages of using microorganisms to make food

Answer 35

➜ microorganisms reproduce quickly so selective breeding quicker
➜ microoganisms can be grown on waste substrates (e.g whey - waste product of cheese making)
➜ growth not seasonal
➜ inputs = cheap like glucose, oxygen, ammonia
➜ fermenters can be set up anywhere in world so food can be produced in extreme climates
➜ low fat, high protein products (mycoprotein)
➜ microorganisms efficient at converting energy into biomass

Question 36

Disadvantages of using microorganisms to make food

Answer 36

➜ sterile technique is expensive
➜ micro reproduce quick so could mutate quick too
➜ perception that food is nutritious but has no flavour etc
➜ bacteria can be infected by viruses
➜ people may not like their food going through microorganisms and wont buy it
➜ fermenter contaminated by pther bacteria could ruin product
➜ fermenter produced products are high in nucleic acids which is harmful in large quantities so it must be processed to be safe to eat

Question 37

Culturing Microorganisms

Answer 37

➜ work aseptically - keep bunsen lit at yellow as flame creates convection current above bench, preventing contamination from air
➜ hot agar jelly poured into sterile petrie dish and left to cool
➜ inoculating loop passed through flame
➜ use loop to take sample from culture tube (flame before and after removing and replacing plug)
➜ open petri teeeny weeeny bit
➜ and wipe on surface of agar
➜ sterilise loop again
➜ tape at intervals around dish and store upside down
➜ incubate 25°C for 24 hrs

Question 38

Batch fermentation

Answer 38

➜ microo grown in batches in fermentation vessel
➜ product removed after culture cycle and fermenter is cleaned for new batch
➜ known as closed culture

Question 39

Continuous fermentation

Answer 39

➜ microo continually grown and the products harvested
➜ nutrients added ans waste is removed through process

Question 40

optimum conditions in fermenter

Answer 40

➜ pH - for enzymes for max rate for max yield
➜ temp - use a water jacket for max yield
➜ O2 - to max resp
➜ nutrient supply - for max yield
➜ agitation - paddles for even distribution of temp, pH, O2 and nutrients
➜ contamination - steam cleaned in between b atches
➜ waste removal - to prevent negative effects on growth

Question 41

Measuring populations of microo

Direct counting

Answer 41

includes all cells (both living and dead) and involves taking samples to count these individual microorganisms

Question 42

Measuring populations of microo

Viable counting

Answer 42

involves culturing samples of microorganisms and counting the colonies that grow (only live cells)

Question 43

Measuring populations of microo

Turbidity

Answer 43

➜ measure of living and dead microorganisms
➜ measure of the cloudiness of a suspension using a colorimeter
➜ as microo can be grown in broth culture
➜ measuring turbidity can be then used to estimate n.o of cells in broth
➜ as microo reproduce suspension becomes more turbid (cloudy)
➜ can be measured by colorimeter and data logger or light sensor or turbidity meter
➜ can then plot a pop curve

Question 44

Standard population growth curves

Answer 44

➜ Lag Phase - pop increases slowly as microo pop adjusts to new env and starts to reproduce
➜ Log Phase - pop grows at exponential rate due to high nutrients
➜ Stationary Phase - pop reaches max as it is limited by resources, n.o of death = n.o being reproduced by binary fiss
➜ Decline Phase - due to lack of nutrients and death due to toxic substance build up, death rate exceeds reproduction rate

Question 45

binary fission

Answer 45

➜ single circular DNA molecule undergoes DNA replication
➜ plasmids present undergo DNA replication
➜ parent cell divides into two cells, with the cytoplasm roughly halved between the two daughter cells
➜ two daughter cells each contain a single copy of the circular DNA molecule and a variable number of plasmids

Question 46

rate of cell division by binary fission:

N = N₀ x 2ⁿ

Answer 46

N = the final number of bacteria
N₀ = the initial number of bacteria
n = the number of division

Question 47

Immobilised Enzymes

(IE)

Answer 47

an enzyme that is attached to an insoluble material to prevent mixing with the product
➜ can be immobolised several ways:
∘ attached to an inert substance - by covalent bonding or attachment
∘ enclosed in a capsule - matrix?
∘ contained within partially perm membr

Question 48

why immobolise enzyme?

Answer 48

➜ for use in industrial processes, enzyme can be reused in future processes rather than being discarded after it has been used once
➜ Reusing the enzyme also avoids the need to separate the enzyme from the product in downstream processing
∘ IE are containted within a column through which substrate is filtered in solution
∘ as substrate runs through column, enzyme substrate complexes are formed
∘ products are produced and these flow out column leaving enzymes behind

Question 49

Advantages of immobilised enzymes

Answer 49

➜ no enzyme in product (uncontaminated) so no need to further filter end product
➜ IE can be reused multiple times - cost effective and efficient
➜ IE has high temp and pH change tolerance
∘ immobilisation makes enzymes more stable

Question 50

Disadvantages of immobilised enzymes

Answer 50

➜ expensive equipment
➜ IE is costly to buy so not helpful for small industries
➜ ROR is sometimes lower when using IE as enzymes can’t freely mix with substrate

Question 51

Lactose free dairy products like milk

Answer 51

∘ enzyme: Lactase
∘ converts lactose to glucose and galactose

Question 52

Semi-synthetic penicillin

Answer 52

➜ overcomes issues of penicillin resistance
∘ Enzyme: Penicillin acylase
∘ converts penicillin to stronger one

Question 53

Glucose products used to sweeten and thicken foods

Answer 53

∘ Enzyme: Glucoamylase
∘ starch and other dextrins to glucose

Question 54

Fructose to sweeten foods that must have low sugar

Answer 54

∘ Enzyme: Glucose isomerase
∘ starch and other dextrins to glucose

Question 55

Purified samples of L-amino acids used in food production

Answer 55

∘ Enzyme: Aminoacylase
∘ Separates out L-amino acids from D-amino acids

Question 56

Acrylamide required in disposable nappy production

Answer 56

∘ Enzyme: Nitrilase
∘ Converts acrylonitrile into acrylamide

Question 57

Using IE to breakdown lactose in milk

Answer 57

➜ there is container of alignate beads bound with lactase
➜ milk is poured in
➜ lactase breaks lactose down into glucose and galactose which can be easily absorbed by the lactose intolerant
➜ lactose = disaccharide