What is
plant tissue culture
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Gottlieb Haberlandt
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F.C. Steward
Plant tissue culture techniques
Explant
It is a plant cell tissue or
an organ which is used to initiate a culture. Practically any part of the plant
can be used as an explant. Explants range in size from 0.1mm to 2 cm.
Points to remember in selecting an explant
1.The explants should be
taken from the best plant
2.Select explants from
plants grown in greenhouses. They have lesser chances of contamination
3.The bigger the explants
the better the response
4.The younger the explants
the better the response
Surface sterilization of the explants
The outer surface of the
explant is infested with microbes. They have to be removed before explants can
be placed on the culture medium. This process is called surface sterilization.
Explants like inner cabbage leaves do not need surface sterilization. One of
the following chemicals can be used.
1.70% ethanol or isopropanol
2.1-10% sodium or calcium
hypochlorite
3.0.1-1% Mercuric chloride
From my personal experience
sodium hypochlorite works best in temperate climate ( it worked well in my
experiments done in USA) but in a tropical climate like that in India 0.1%
mercuric chloride is a better choice.
Because of the presence of a
waxy layer on the explant a surfactant ( a chemical which removes surface
tension of water) is used in the surface sterilizing solution. A drop of liquid
soap or Tween 20 can be used.
Procedure for surface sterilization
1.Wash explants in running
water for 30 minutes
2.Treat with 0.1% mercuric
chloride containing surfactant
3.Wash thrice in sterile
distilled water
4.Trim on sterile filter
paper in sterile petridish
5. Inoculate on the medium
Steps 2-5 should be done in
a laminar flow hood / clean air work station
If the explant turns brown
during treatment dip in an antioxidant solution before trimming. An antioxidant
solution is made by dissolving 100mg of ascorbic acid and 150 mg of citric acid
in a litre of water. Sterilize before use
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