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Year 1 Biology Ellie M > Biotechnology > Flashcards

Biotechnology Flashcards

1
Q

When was the first recombinant organism produced?

A

E. coli in 1972

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2
Q

What is white biotechnology?

A 
Industrial biotechnology- the use of living organisms or their derivatives to make industrial products
Chemicals 
Amino acids
Vitamins 
Enzymes
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3
Q

History of citric acid production

A

Produced in the UK from 1826
Calcium citrate produced from lemons and converted to citric acid chemically
1917 - Aspergillus niger (fungus) found to produce copious amounts of citric acid
1923 - large scale production began in New York
Over 1-2 million tonnes produced worldwide

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4
Q

How is acetic acid produced?

A

Fermentation of ethanol or methanol by microbes

200,000 tonnes produced annually

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5
Q

How is butanol produced?

A

From petroleum for fermentation of Clostridium acetobutylicum
Used in plastics, paint, resins, etc
1.2 million tonnes produced annually
Byproduct is acetone

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6
Q

How is lactic acid produced?

A

Half by microbes, the rest is chemical

Used as acidified, preservative and in plastics

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7
Q

Where were enzymes traditionally obtained from?

A

Microorganisms, plants and animals

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8
Q

How are enzymes produced now?

A 
From fungi
Submerged cultures (in liquid) of Aspergillus oryzae used to produce enzymes in large bioreactors
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9
Q

Advantages of obtaining enzymes from fungi

A

Cheap
High yields
Continuous

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10
Q

Uses of pectinases

A

Break down peptin in manufacture of fruit juice and baby food

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11
Q

Uses of proteases

A

Leather tanning

Many other uses

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12
Q

Uses of phytases

A

Added to animal feed to enable digestion of phosphate

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13
Q

What enzymes are used in detergent?

A

Modified subtilisin from Bacillus subtilis
Alkaline proteases
Amylase
Lipases

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14
Q

What are the benefits of immobilising enzymes?

A 
Enzymes can be recycled
Preserves expensive enzymes - lower production costs
Absent from end product
Increased stability
Bound to gel or membrane
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15
Q

What does glucose isomerase do?

A

Converts glucose to fructose

High fructose corn syrup went into mass production in the 1960s

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16
Q

How has immobilisation changed the use of glucose isomerase?

A

Before the syrup only contained 15% fructose because enzyme too expensive to produce
Afterwards fructose yield was 42%
Still cheaper to produce sucrose
Technological advances in 1970s reduced glucose isomerase production costs, and sucrose prices rose, so HFCS now economically viable

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17
Q

How much is the production cost of glucose isomerase reduced by due to immobilisation?

A

40%

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18
Q

What is the current fructose content of high fructose corn syrup?

A

55%

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19
Q

What is red biotechnology?

A 
Health-related biotechnology
Biopharmaceuticals
Recombinant proteins
Vaccines
Stem cells
Animal models
Gene therapy
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20
Q

Recombinant proteins

A

Over 100 in use, on which 50 are antibodies ($50 billion)

Main uses are replacement for missing/defective proteins and inhibition of infectious agents

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21
Q

Examples of recombinant proteins

A

Insulin
Antibodies
Blood clotting factors
Vaccines

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22
Q

What was the first commercially available GM hormone?

A

Insulin - lowers blood glucose levels

Produced by E. coli

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23
Q

What methods have overcome problems in insulin production?

A

Allergic reactions to non-human insulin
Had to tweak sequence using enzymes to modify the protein
Also tweaked DNA sequence to prevent clumping when being injected

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24
Q

How much are vaccines worth?

A

$40 billion per annum

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25
Q

What was the first vaccine?

A

Developed by Edward Jenner in 1796
Live cowpox virus infection protected against smallpox virus
First infectious disease to be eradicated worldwide (1977)

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26
Q

What is a subunit vaccine?

A

Fragments of the pathogen e.g. viral coat proteins or lipids
Developed prior to recombinant DNA technology
Hepatitis B vaccine was first example

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27
Q

How was the hepatitis B vaccine developed?

A

Blood-borne virus that causes liver disease

The virus was isolated from infected blood and the proteins purified and used in the vaccine

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28
Q

How are subunit vaccines produced now?

A

The relevant protein-encoding genes are cloned into plasmids, transformed into yeast or a cell line and the proteins would be purified

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29
Q

What is an inactivated vaccine?

A

Killed pathogen
Most common type
Vaccinate against polio (Sulk vaccine), rabies, influenza, hepatitis A
Many pathogens cannot be isolated or cultured in vitro or are too expensive to culture
Risk of infection to biotech workers

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30
Q

What are attenuated vaccines?

A

Live, weakened pathogens that no longer express the toxin gene
Can be a natural mutant or a GM mutant
Could be a related virus
Vaccinate against polio (Sabin vaccine), MMR, tuberculosis, chickenpox, cholera
Safer to produce, but needs a lot of research to identify the toxic gene
Risk that attenuated pathogens may revert to pathogenic strain

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31
Q

How are new vaccines developed? (first method)

A
  1. Genome sequenced to find antigens
    Identify genes and clone into expression library
    Determine which proteins are responsible for the immune response
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32
Q

How are new vaccines developed? (second method)

A
  1. Use viral genomes
    Can use Vaccinia virus to make new vaccines
    Clone genes of interest into a plasmid and insert into vaccine genome
    Use recombinant Vaccinia virus as a vaccine against smallpox as well as other illnesses
    Cheaper
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33
Q

How are new vaccines developed? (third method)

A
  1. DNA-based vaccines
    Add the gene encoding the antigen into a plasmid
    Bind the DNA to a charged particle and inject it
    The DNA will bind with genomic DNA particles and the antigen will be expressed temporarily, triggering a localised immune response
    Cheaper to make and easier to store
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34
Q

When and what was the first DNA-based vaccine?

A

2005

Produced against the West Nile Virus

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35
Q

How are new vaccines developed? (fourth method)

A
  1. Edible vaccines
    Antigens can be expressed on plants, then they can be eaten
    However lots must be eaten and it must be raw
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36
Q

What are stem cells?

A

Undifferentiated cells, which can differentiate into specialised cells and replicate to make more stem cells

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37
Q

What are the two types of stem cells?

A

Adult stem cells

Embryonic stem cells

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38
Q

What are totipotent stem cells?

A

They can regenerate into all cell lineages to regenerate a whole organism e.g. in plants and fungi, or embryonic stem cells within the first couple of divisions (they can generate both embryonic and extra-embryonic cells)

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39
Q

What are pluripotent cells?

A

Capable of forming all the cell lineages within an embryo, but not extra embryonic lineages

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40
Q

What are multipotent cells?

A

Have the potential to differentiate into many, but not all, cell lineages

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41
Q

What are hematopoietic stem cells (HSCs)

A

Found in bone marrow

Multipotent - replenish red and white blood cells

42
Q

What are the two places that hematopoietic stem cells are found?

A
  1. Vascular niche - near blood vessels within the marrow

2. Endosteal niche - at the interface between the bone and marrow

43
Q

What two cell lines do hematopoietic stem cells differentiate into?

A
  1. Lymphoid progenitor - differentiate into T and B cells

2. Myeloid progenitor - differentiate into red blood cells, macrophages and neutrophils

44
Q

What are intestinal epithelial stem cells (ISCs)?

A

Found in the small intestine
Also called crypt base columnar cells (CBCs)
Can differentiate into four cell types

45
Q

What are the four cell types that CBCs can differentiate into?

A

Absorptive epithelial cells (enterocytes)
Goblet cells - secretes main component of mucus
Enteroendocrine cells - control glucose levels, food intake and stomach emptying
Paneth cells - secrete anti-microbial compounds

46
Q

What are embryonic stem cells (ESCs)?

A

Derived from the blastocyst
Pluripotent, so capable of forming all the cell lineages within an embryo, but not extraembryonic lineages
Cells can be grown in culture (since 1998)

47
Q

What are induced pluripotent stem cells (iPSCs)?

A

Adult cells the have been undifferentiated back to stem cells
Changes in gene expression allow reversion of adult cells into stem cells
Less controversial than embryonic stem cells
Not clear if identical to ESCs
Low efficiency of production
Can cause rumours
STAP cell comtrovery

48
Q

What is the controversy over STAP cells?

A

Research on generating pluripotent stem cells by subjecting ordinary cells to certain types of stress (e.g. bacterial toxins or physical trauma) was published in 2014 by Haruko Obokata. However, others were unable to replicate it - may be error or fraud

49
Q

What stem cells are often used in stem cell therapy?

A

Hematopoietic stem cells most commonly used (bone marrow transplant)
HSCs also found in umbilical cord blood and some circulate in blood

50
Q

What could stem cell therapy be used to treat?

A 
Leukaemia
Sickle cell anaemia
Tissue regeneration
Reverse ageing
Parkinson’s
Diabetes
Spinal cord injuries
Replacement for animal testing
51
Q

Treatment for diabetes

A

Induce stem cells to differentiate into pancreatic B cells that produce insulin
Use these to treat type 1 diabetics (who cannot produce insulin)
Tested in mouse models and mice have been shown to produce insulin
More work needed to be done to prevent immune system attacking B cells

52
Q

How to transform embryonic stem cells

A
  1. Make vector containing gene if interest
  2. Transform stem cells
  3. Insert stem cells into embryo
  4. Implant into mouse
53
Q

How are knockout mice produced?

A
  1. Clone regions flanking gene into a vector
  2. Introduce DNA construct into mouse
  3. Construct replaces or disrupts gene, so no protein is made and gene function can be analysed
  4. Used in research of human diseases
54
Q

What are knockout mice?

A

They have genes ‘knocked-out’ to analyse their function

55
Q

How is cloning achieved?

A
  1. Remove nucleus from egg cell
  2. Add a different nucleus
  3. Allow to develop into frog
56
Q

What did cloning show?

A

The the nucleus of a differentiated cell can regenerate an entire organism

57
Q

What is forensic biotechnology?

A 
Use of science in gathering evidence
Fingerprints
DNA technology
Blood type
DNA fingerprinting
DNA databases/barcoding
Biometrics (body measurements and calculations)
58
Q

Features of bloodtyping

A 
Glycolipid antigens A, B and O
Different alleles of the same gene
O has one fewer carbohydrate
A has N-acetyl-galactosamine
B has galactose
59
Q

What is DNA fingerprinting?

A

Analysis of DNA fragments for identification
Developed in 1980s
Early method used restriction fragment length polymorphisms (RFLPs)

60
Q

Process of early DNA fingerprinting

A
  1. DNA was extracted, cut with restriction enzymes and run on gel
  2. A Southern blot was used, where DNA was transferred to a nylon filter
  3. Radioactively labelled DNA probe were used, which bound to complementary DNA
  4. An autoradiograph is generated, where radiation-sensitive film is placed over the blot and DNA bound to the probes appear as bands on the film
  5. The restriction pattern is matched to DNA found on the victim
61
Q

Current process of DNA fingerprinting

A

DNA is now amplified by PCR and sequenced
Mini satellites and microsatellites often used
Can be run on a gel or visualised by qPCR or sequencing software

62
Q

What is a minisatellite?

A

Variable number tandem repeats (VNTRs)
10-100 base pairs repeating 5 to 50 times
Segment of repetitive DNA

63
Q

What is a microsatellite?

A

Short tandem repeats (STRs)

2-5 base pairs repeating 5 to 50 times

64
Q

Examples of phenotypic biometrics

A

Fingerprints
Retinal scans - look at the pattern of blood vessels behind retina
Iris recognition systems
Facial recognition

65
Q

What is DNA barcoding?

A

Used to identify species
Uses conserved regions of genome (e.g. chloroplasts in plants, mitochondria COI in animals)
Can be used to analyse what is in food and to ID insect larvae

66
Q

What is bioremediation?

A

The use of living organisms or their products to break down waste and pollutants in the environment
Can clean up organic waste, chemical spills, pesticides, heavy metals, etc.
Uses natural processes and reactions

67
Q

How are organisms obtained for bioremediation?

A

Can search for organisms that degrade waste and use them directly
Can genetically modify other organisms to express the relevant genes

68
Q

What are some common chemical pollutants?

A 
Benzene
Chromium
Naphthalene
Radioactive compounds
Trace metals
Trinitrotoluene (TNT)
69
Q

Describe bioremediation of ground water

A

If drinking water is contaminated, need a bioreactor containing bacteria to separate the clean water

70
Q

What microbes are used for bioremediation?

A
  1. Petroleum-eating bacteria: isolated strains of Pseudomonas found in contaminated soils contained plasmids encoding genes for degrading organic compounds such as naphthalene, octane and xylene
  2. E. coli used for heavy metals
  3. Metallothioneins (proteins on cell surface)to neutralise heavy metal pollutants such as cadmium and mercury
  4. Fungi Phanerochaete spp. can degrade toxic chemicals such as creosote and other pollutants that degrade bacteria
  5. Fusarium oxysporum and Mortierella hyaline can degrade asbestos and heavy metals
71
Q

Describe the Exxon Valdez disaster

A

1989
Oil tanker ran aground and released 42 million litres of oil off Alaskan coast
Oil will remain for hundreds of years

72
Q

What are steps for bioremediation of oil spill?

A
  1. Clear oil from surface using skimmers and vacuums
  2. Wash rocks with water
  3. Added fertiliser to encourage growth of bacteria to degrade o
73
Q

Describe the Deepwater Horizon oil spill

A

2010
Explosion released 600 million litres of oil into Gulf of Mexico
Oil removed by various methods
Bioremediation degrades 50% of oil released

74
Q

What types of bacteria are used to removed the sulphur from fossil fuels and reduce their toxicity?

A

Thiobacillus and Sulfolobus can be used to convert inorganic FeS2 to sulphates that can be washed away
That accounts for 30% of sulphur in fossil fuels, the rest is in thiophene rings
Rhodococcus can break down thiophene but is hard to culture
Transformed E. coli may be able to break down thiophene

75
Q

Describe bioremediation of lignocellulose

A

It is a waste agricultural and industrial product
Microorganisms can degrade cellulose and hemicellulose, but not lignin
White rot fungi can degrade lignin using peroxidases to break the bonds between phenols in lignin

76
Q

What is a white rot fungi that can degrade lignin?

A

Phanerochaete chrysosporium

77
Q

Using bioremediation to recover valuable metals

A

Cooler, nickel, boron, gold
Many microbes convert metal products into metal oxides or ores
Useful for recovery of metals from waste solutions from industrial manufacturing processes

78
Q

Bioremediation of radioactive wastes

A

Most radioactive materials kill microbes, but some strains have a potential for degrading radioactive chemicals
No bacterium so far can completely metabolise radioactive elements into harmless products

79
Q

What is phytoremediation?

A

Using plants for bioremediation of soil, water and air
Around 350 plant species naturally take up toxic materials
Poplar, juniper, grasses, Agrostis
Sunflower plants removed radioactive cesium and strontium from Chernobyl
Water hyacinths removed arsenic from water supplies in Bangladesh and India

80
Q

What is biofuel?

A

Fuel produced through biological processes

81
Q

What is the total biofuel production in billions of litres?

A

81 billion litres per year

82
Q

Issues with biofuel production

A 
Expensive
Subsidy dependent
Fuel instead of food
What to do with waste products
Impacts on biodiversity
83
Q

Biogas

A

2 million people worldwide burn biogas for fuel

Could produce gas from animal dung in small bioreactors

84
Q

Bioethanol

A

Sugarcane or maize often raw materials
Microbes convert sugars into ethanol
Production is expensive
Fuel additive

85
Q

Biodiesel

A

Derived from plant oils and animal fats

Standalone fuel - replacement for diesel

86
Q

Lignocellulose biomass as a biofuel

A

Agricultural waste products - woody crops, sawdust
Do not need much water or fertiliser to grow
More sustainable
Still for biodiesel and bioethanol
Less competitive due to difficulties degrading lignocellulose

87
Q

Biofuels from algae and cyanobacteria

A

Still under development
No competition for land use
Oils easily refined into diesel
Easy to genetically manipulate algae into ethanol and butanol
Can also make biogases such as biohydrkgen and biomethane
Algae produce 10x the output of traditional biofuel feedstocks
Lots of fertiliser needed

88
Q

What % of fuel in the UK is biofuel?

A

3%

89
Q

What greenhouse saving do biofuels have compared to fossil fuels?

A

80%

90
Q

Bioremediation of paper waste

A

Paper is mainly cellulose
Various microbes produce different cellulases
These cellulases have been purified and shown to degrade paper to glucose
Glucose could then be used to produce ethanol
Inefficient multi step process
Cellobiose acts as feedback inhibitor of cellulose degradation
Glucose inhibits hydrolysis of cellobiose
End products of this reaction therefore have to be removed quickly to allow continuous degradation

91
Q

Fuel from microbes

A

Derived from plant oils and animal fats - could otherwise be used for food
Engineer E. coli to produce large quantities of fatty acids

92
Q

What is green biotechnology

A

Use of crop plants and agricultural systems
Humans have modified plants for thousands of years
Since 1920s there has been mutational breeding, plant propagation, etc
Genetic modification

93
Q

What is Agrobacterium tumefaciens?

A

Plant pathogen used in molecular biology
Causes crown gall (1907)
Induced plant to produce excess auxin and cytokinin
Galls contain opines synthesised by genes on the Ti plasmid

94
Q

Other plant transformations

A

Gene gun or particle bombardment

Gold particles are coated in the DNA construct to be introduced, then they are fired at target cells

95
Q

GM crops

A

Also called biotech or transgenic crops
First commercial GM crops in 1994
Grown in 26 countries
$18 billion annually

96
Q

What 4 GM crops dominate global production?

A

Soybean (50%)
Maize (31%)
Cotton (13%)
Oilseed rape (4%)

97
Q

Two main traits of GM crops

A

Herbicide tolerance

Insect resistance

98
Q

Resistance to glyphosate

A

Herbicide
Blocks amino acid synthesis
Introduce a mutant EPSPS gene from Agrobacterium tumefaciens (aroA); glyphosate cannot bind
Plant is resistant to glyphosate

99
Q

GM plants producing Bt toxin

A

cry genes from soil bacterium Bacillus thuringiensis
Toxic protein, which solubilises when ingested by insects
Binds to epithelial cell’s within the digestive tract and creates holes
Insects die within a few days
Spray plants with it in organic agriculture

100
Q

Other GM traits

A

Herbicide tolerance
Virus resistance - mosaic viruses
Quality traits - drought tolerance, golden rice, photosynthetic genes

Year 1 Biology Ellie M (28 decks)

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