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Molecular Biology Flashcards > Genetic Engineering of Plants > Flashcards

Genetic Engineering of Plants Flashcards

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1
Q

Learning Outcomes

A

Students will be able to….
* explain why all domesticated plants are genetically modified and recognise
that some plants are naturally occurring transgenic plants.
* describe the properties of Agrobacterium tumefaciens and it’s Ti plasmid and
relate those to infection and transformation of plants.
* compare & contrast procedures & workings of plant transformation methods.
* define terms such as: transgene, sense and antisense transgenes and relate
those to effects they have on protein expression.
* describe examples of sense and antisense transgenic plants and the effects
the genetic manipulations have on the plant.
* discuss potential benefits of transgenic plants

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2
Q
A
  • A single amino acid substitution – so a single mutation
    in the DNA - leads to the abolishment of the seed coats
    in Teosinte and the naked kernels found in maize
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2
Q

Limitations of traditional breeding methods

A

➢ Interbreeding limitations: only closely related species can interbreed
➢ Lack of desired traits: may not exist in the plant or its close relatives
➢ Time - life cycles often longer than a year
➢ Undesirable traits in close relatives - uncontrolled inheritance of undesirable traits
along with the desired ones

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2
Q

Genetically Modified Plants

A

All domesticated plants are genetically modified by selection and breeding for desired traits, e.g., disease resistance, showy flowers, large fruits

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2
Q

What are those transgenic thingies?

A
  • Transgenic technology - insertion of transgene(s) into the
    genome of an organism using recombinant DNA techniques
  • Transgene - a gene that has been transferred naturally, or by
    genetic engineering techniques, from one organism to another.
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3
Q
A

─ T-DNA is cut out of Ti plasmid
at repeat (R) regions by Vir
encoded proteins
─ Vir proteins ensure that T-DNA
is transferred to plant nucleus
─ integrated into plant
chromosome

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3
Q

Using Agrobacterium to produce transgenic Plants

A

Ti (tumour-inducing) plasmid
* T-DNA = transfer DNA
* vir - essential for T-DNA transfer
(virulence)
* R - repeated regions
* onc - encodes p

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4
Q

Agrobacterium tumefaciens

A
  • soil bacterium, gram
    negative
  • naturally infects diverse
    plants at wounded sites
  • causative agent of crown
    gall = tumour/
    undifferentiated cells
  • used by plant molecular
    biologists to transform
    plants, e.g., introduce a
    transgene
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5
Q

Separating virulence factors from the T-DNA

A
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5
Q
A
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6
Q

Separating virulence factors from the T-DNA

A

Binary vector system: two Ti plasmids
➢ Plasmid 1: no vir genes, has T-DNA, researchers insert transgenes into T-DNA
➢ Plasmid 2: no T-DNA, has vir genes
▪ both plasmids must be present in Agrobacterium for transgene insertion

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6
Q

Transforming plants with Agrobacteria using the
binary vector system

A

➢ T-DNA plus transgene
on plasmid 1 is excised
by virulence proteins
encoded by the
plasmid 2
➢ Just as in nature, TDNA is transferred into
plant nucleus by vir
proteins and integrated
into chromosomes
(randomly)

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7
Q

Transforming plants with Agrobacteria using the
binary vector system

A

Genes added to T-DNA by researchers:
➢ selectable marker: e.g., antibiotic resistance gene
➢ Gene of interest: new trait encoding gene, gene
encoding a protein to be studied etc.

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8
Q

Transforming plants with Agrobacteria using the
binary vector system

A
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8
Q

The technical part of plant transformation using
Agrobacterium

A
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9
Q

An Agrobacterium-free plant transformation system

A
  • Biolistic- or microprojectile
    bombardment-mediated
    transformation
  • Transgene-coated gold particles
    (“bullets”) are “shot” into plant tissue.
  • DNA is incorporated into plant
    chromosome
  • After “shooting”, plants are
    regenerated as described on slide
    ‘The technical part of plant
    transformation using Agrobacterium’
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9
Q

The technical part of plant transformation using
Agrobacterium

A
  • Incubate leaf discs with genetically engineered Agrobacterium
  • Place leaf discs on selective medium (e.g. with antibiotic) that also contains plant
    hormones ➔ only transgenic plant cells survive and form a callus = undifferentiated cells
  • Use different plant hormones to induce shoot & root formation
10
Q

An Agrobacterium-free plant transformation system

A

Advantages:

  • no Agrobacteria needed
  • can be used for plants that cannot be
    transformed with Agrobacteria
10
Q

An Agrobacterium-free plant transformation system

A

Disadvantage:

  • requires callus formation and
    regeneration of plants
11
Q

The technical part of plant transformation using
Agrobacterium

A
12
Q

An Agrobacterium-free plant transformation system

A
13
Q

A variation of plant transformation using Agrobacterium – Floral Dip transformation

A
13
Q

Transgenic sense and antisense plants

A
14
Q

A variation of plant transformation using Agrobacterium – Floral Dip transformation

A
  • The transfer of T-DNA culture
    from Agrobacterium to
    plant nucleus works as
    described on slide
    ‘Transforming plants with
    Agrobacteria’
  • Advantage: no callus
    formation needed
  • Problem: does not work
    with all plants
14
Q

Sense plants: Insecticide

A
15
Q

Sense plants: Herbicide Resistance

A
  • Roundup Ready® cotton, soybean, canola
  • Roundup® - broad spectrum herbicide
  • active ingredient - glyphosate - inhibits enzyme in synthesis of some amino
    acids
  • Roundup Ready® crops contain modified enzyme not recognized by glyphosate
15
Q

Antisense plants:
Foods with Enhanced Flavour and Shelf-life

A
  • FlavrSavr Tomato
  • antisense for a cell wall degrading enzyme
  • degradation of cell walls slowed ➔ increased time on the vine which increases flavour,
    improved handling of fruit during transport and longer shelf life (tomatoes shown were
    on the shelf for four month)
  • Failed: experiment worked but company did not get the marketing correct
16
Q

Sense plants:
Transgenic Plants as Bioreactors

A

Generation of antibodies, vaccines, medical and industrial proteins

Transgenes for the production of
biodegradable plastics

  • pink dots and areas are granules of the plastic
16
Q

Sense and antisense:
Genetically Modified Ornamentals

A
17
Q

Natural Transgenic Plants

A

Cultivated sweet potato
* first food crop found with T-DNA
(2015)

  • Not found in wild relatives ➔
    suggests genes in the T-DNA
    code for desirable traits that
    were selected for over millennia

Other plants with T-DNA
* wild tobacco species
* Linaria
Will this change public
perception of transgenic crop
plants?

18
Q

Putting food and fodder security into perspective

A

In 2012 (2018):
* 8.2 (9.6) million people worldwide died from cancer (World Health
Organization)
* 3.1 (6.2) million children under the age of 5 (15) died of malnutrition and
hunger-related diseases in developing countries (United Nations
Children’s Fund) …and how many people over the age of 5 (15)?
* Total deaths from hunger = 9 million

18
Q

United Nations Food and Agriculture Organization
(FAO) predicts:

A

▪ Requires new technologies and improved practices
* smarter irrigation methods – e.g. soil moisture monitoring
* sustainable farming practices – e.g. inter-planting with legumes
* targeted breeding – BOTH traditional methods and biotechnology

18
Q

Should We Consider Transgenic Crops?

A
  • Production of grains is not keeping pace with human and animal
    consumption
  • Grain reserves are at historical lows
18
Q

United Nations Food and Agriculture Organization
(FAO) predicts:

A

➢ 70% increase in world food production required to meet demands of 2.3 billion
more people in 2050

19
Q

Should We Consider Transgenic Crops?

A
  • global population growth, especially in developing countries
  • loss of agricultural land due to degradation and increasing urbanisation
  • climate change
  • limitations on fertilizers
  • limitations of water
  • demands for biofuels
19
Q

United Nations Food and Agriculture Organization
(FAO) predicts:

A

▪ Requires responsible practices and understanding
* understanding what transgenes code for and what is consumed
* Canola oil is the part of canola consumed ➔ has no transgenic
DNA
* Transgenic lettuce: eat leaves, digested in mouth, stomach and
guts ➔ transgenic DNA is digested not entered into our cells

20
Q
A

1) Vitamin A deficiency
* affects 250,000 to 500,000 children / year, leads to blindness
* prevalent in developing countries where rice is a staple food
* β-carotene (pro-vitamin A) – synthesised in green tissues of rice, but not the seeds

21
Q
A
  • “Golden Rice 2” – genetically engineered to make β-carotene in seeds
    ➢ transgenes – from corn and a plant pathogen (bacterium)
    ➢ contains recommended daily allowance of vitamin A (form of β-carotene) in 100-
    200 g of rice = typical daily consumption for children in rice-based societies
22
Q
A

2) Micronutrient (e.g. Zn, Fe) and protein deficiencies
* affect over 2 billion people worldwide
* lead to anemia, growth and development problems

22
Q
A
  • wild durum (pasta) wheat contains a gene
  • associated with its higher protein, Zn and Fe content than
    domesticated wheat
  • domesticated wheat varieties have an inactive form of the
    gene, we bred wheat that has lost this gene function.
    Should the wild durum wheat gene be inserted into domesticated wheat?
22
Q

Can you …

A
  • … explain why some plants are naturally occurring transgenic plants and
    why all domesticated plants are genetically modified? .
  • … compare the natural ability of Agrobacterium tumefaciens to infect
    plants with the use of this bacterium to generate transgenic plants?
  • … explain the changes that were made to the Ti plasmid before it could
    be used to generate transgenic plants?
  • … give an overview of two plant transformation methods using
    Agrobacterium and one that does not require Agrobacterium. Can you
    state similarities and differences?
  • … explain the terms transgene, sense and antisense transgenes and
    relate those to effects they have on protein expression using examples
    from the lecture.
  • … give examples of transgenic plants that were created to overcome
    problems and relate this to why transgenic plants may be useful to secure
    food production

Molecular Biology Flashcards (28 decks)

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