Chapter 22 - Cloning and Biotechnology

Natural cloning in plants

• bulbs

  • leaf bases swell with stored food from photosynthesis

  • buds form internally

  • develop in the next growing season

• runners

  • lateral stem grows away from parent plant

  • roots develop where it touches the ground

• rhizomes

  • specialised horizontal stem underground

  • buds develop and form new vertical shoots

• stem tubers

  • tip of an underground stem becomes swollen with stored food

  • buds develop to produce new shoots

• used in horticulture to produce new plants

  • splitting of bulbs

  • cuttings (sections of stems) are planted with rooting hormones

Micropropagation

• making large numbers of genetically identical offspring from a single parent using tissue culture techniques

• used when a desirable plant

  • does not readily produce seeds

  • doesn’t respond well to natural cloning

  • is very rare

  • has been genetically modified or selectively bred

  • is required to be pathogen free

• take a sample of tissue (meristem) and dissect in sterile conditions

• sterilise in bleach etc

• material from the plant is known as explant

• explant placed in sterile culture medium containing plant hormones which stimulate mitosis

• callus is divided up and individual cells are transferred to a new culture medium to stimulate development of plantlets

• plantlets are potted

Advantages and disadvantages of micropropagation

Advantages:

• allows for the rapid production of large numbers of plants with known genetic makeup

• producing disease free plants

• possible to produce viable numbers of plants after genetic modification

• can produce large numbers of seedless plants (sterile)

• provides a way of growing plants which are naturally infertile

• can reliably increase the numbers of rare plants

Disadvantages:

• produces a monoculture

• relatively expensive

• requires skilled workers

• explants and plantlets are vulnerable to infection by moulds

• if the source material is infected with a virus, all the clones will be too

Natural animal cloning

• common in invertebrates

• some can regenerate entire animals from fragments of the original e.g. starfish

• some produce clones as part of normal reproductive process

• females can produce offspring without mating in some insects

Cloning in vertebrates

• monozygotic twins (identical)

  • embryo splits to form 2 separate embryos

• artificial twinning

  • split in early embryo is produced manually

  • can be used to produce a number of identical offspring

  • e.g. in cattle

    • cow treated with hormones to super-ovulate

    • ova fertilised

    • cells are split to produce several smaller embryos

    • grown in the lab for a few days before being implanted in a surrogate

  • somatic cell nuclear transfer

    • nucleus is removed from a somatic cell

    • nucleus removed from a mature ovum harvested from a different female of the same species (enucleated egg cell)

    • nucleus from somatic cell is placed into enucleated egg cell and given an electric shock to fuse them and so it begins to divide

    • developed embryo is transferred into a third animal

    • new animal is a clone of the animal that the somatic cell came from

Pros and cons of animal cloning

Pros:

• enables high yielding animals to produce more offspring

• enables the success of the male passing on desirable genes

• enables genetically modified embryos to be replicated

• enables rare, endangered or extinct animals to be reproduced

Cons:

• inefficient process

• many cloned animals fail to develop

• many cloned animals have shortened lifespans

• has been relatively unsuccessful so far

• ethical concerns

• strict legislation to prevent cloning of humans

Bioremediation

• use of biological systems to remove soil and water pollution

What are the advantages of using of microorganisms?

• no welfare issues

• enormous range of microorganisms capable of carrying out many different chemical syntheses

• genetic engineering allows artificial manipulation of microorganisms

• microorganisms have a short lifecycle

• the nutrient requirements are cheap

• required conditions are cheap - low temperature, oxygen and food etc

How are microorganisms used for medication?

• producing penicillin

  • antibiotic produced by a mould

• making insulin

  • used for type 1 (occasionally type 2) diabetes

• bioremediation

  • microorganisms used to break down pollutants and contaminants in soil or water

How are microorganisms grown?

• inoculating broth

• inoculating agar

Growth of bacterial colonies

• lag phase

  • bacteria are adapting to new environment

  • growing and synthesising the enzymes they need

• log / exponential phase

  • rate of reproduction is close to its theoretical maximum

• stationary phase

  • total growth rate = 0

  • reproduction rate = death rate

• death / decline phase

  • reproduction has almost ceased

  • increasing death rate

What are the limiting factors that prevent bacterial growth?

• nutrients available

• oxygen levels

• temperature

• build up of waste

• change in pH

What are the advantages of using isolated enzymes instead of whole organisms?

• less wasteful as they don’t require substrate to grow / reproduce

• more efficient as they work at much higher concentrations

• more specific - no unwanted enzymes

• ideal conditions can be used

• less downstream processing as other byproducts are not produced

What are the advantages of using extracellular enzymes instead of intracellular?

• extracellular are excreted by microorganisms making them easy to isolate

• only produce relatively few types of extracellular enzymes - easy to identify

• more robust than intracellular

What are the pros and cons of using immobilised enzymes?

Advantages:

• can be reused

• easily separated from reactants and products (reduce downstream processing)

• greater temperature tolerance

• smaller long term costs as they can be reused

• easy to manipulate

Disadvantages:

• reduced efficiency

• higher initial costs of materials

• higher initial costs - bioreactor

• more technical issues

How are enzymes immobilised?

• surface immobilisation

  • bound to the surface of insoluble supporting materials either by adsorption or covalent or ionic bonds

  • adsorption

    • simple & cheap

    • enzymes can be lost relatively easily

  • covalent / ionic bond

    • strongly bound

    • active site may be modified

• entrapped in a matrix

  • widely applicable to different processes

  • expensive & difficult to entrap

• encapsulated in a microcapsule

  • simple to do

  • relatively expensive

• behind a semi permeable membrane

What are the types of bioprocesses for culturing microorganisms?

• batch fermentation

  • microorganisms are inoculated into a fixed volume of medium

  • as they grow, nutrients are used up

  • biomass and waste products build up

  • during stationary phase - form desired end products

  • stopped before death phase

• continuous culture

  • microorganisms inoculated into sterile nutrient medium

  • nutrients added continually

  • culture broth is continually removed