Transcript Structure, function and growth of prokaryote and eukaryote

Structure, function and growth of
prokaryote and eukaryote cells
(iii) Differentiation of cells
into tissues and organs.
(iv) Cell and tissue culture
–Mammalian
cell cultures
–Bacterial and Fungal
Cultures
(v) Plant tissue culture
Basic requirements for cell culture






Source of cell material
A suitable container for cell growth
Growth medium containing essential
nutrients
Opportunity for gas exchange
Control of abiotic factors e.g. temperature
and pH
Avoidance of contamination
Aseptic techniques

Methods employed to prevent contamination
–
E.g. sterilisation of equipment, media (substance
in which the cells grow) and surfaces



–
Autoclaving = heating substance to a high temperature
(usually 121oC) under high pressure to kill microbes and
their spores.
Swabbing with disinfectant or alcohol
Running equipment through a flame before use.
Minimising exposure of equipment to the air
Aseptic Techniques cont….

Biological Safety
cabinets
Aseptic techniques cont…
Culturing Mammalian Cells

Source of Cell material
–
–
–
Tissue samples are treated with a proteolytic
(protein digesting) enzyme e.g trypsin to separate
cells from each other
Normal cells die after a finite number of divisions
when grown in culture, making long-term culturing
difficult.
For this reason cell lines are often prepared from
cancer cells or cells that have been genetically
modified to become immortal.
Culturing Mammalian Cells cont…

Suitable container
–
Mammalian cells require a solid surface for growth in
culture
Culturing Mammalian Cells cont…

Growth medium
– Is complex
– Minimum growth medium contains balanced salt
solution, amino acids, vitamins and glucose.
– Additions are made to minimal growth media
according to the requirements of the cells.
– Antibiotics may be added to prevent
contamination
– Foetal Bovine Serum (FBS) (or other animal
serum) is essential to promote cell proliferation
(division)
Culturing Mammalian Cells cont…

Events in culturing mammalian cells
–
–
–
–
Growth medium is inoculated with cell source
Cells adhere to the surface of the container
Cells spread out and then divide to form a
monolayer
When the entire surface is covered with cells just
touching one another, they are said to be
confluent or to have reached confluence.
Culturing Mammalian Cells cont…
Cells spreading out and dividing
Confluent cells
Note: If a single cell was isolated and used to inoculate the culture, then
all of these cells will be clones of one another.
Culturing Bacteria and Fungi

Source of Cell Material
–
–

Commercially bought
Isolated from environment
Suitable Container
–
Bacteria and liquid can be grown in solid or liquid
medium and do not need a solid surface to
adhere to.
Culturing Bacteria and Fungi cont.
Culturing Bacteria and Fungi cont.

Growth Medium
–
–
–
Is basic
Most microbes only require a source of carbon
(usually a sugar for energy and to make basic cell
components)
Some require special additives
Culturing Bacteria and Fungi cont.

Events in growing Bacteria and Fungi
–
–
–
After an initial lag phase microbes will undergo
exponential growth.
Cultures then enter a stationary phase as
nutrients become limited and wastes accumulate.
Death will eventually occur if not sub-cultured
Culturing Bacteria and Fungi cont.
Learning Activities




Read DART pg 20 – 22
Scholar (Unit 1, 3.1 – 3.3)
Techniques in Microbial Culture
Cloning cells for an Interferon Bioassay