Nexus Research Group - Tissue Culture

Tissue Culture

Tissue culture involves getting cells to grow in a petri dish instead of as part of a whole organism.

Many types of plant, such as radiata pine, can be grown from in petri dishes from clumps of cells instead of from seeds...

Method:

Keeping conditions sterile is of the utmost importance as micro-organisms (bacteria, moulds and viruses) can contaminate the petri dish and kill of the cells you are trying to grow...

Obtain an African Violet leaf. Sit a scalpel and a pair of tweezers in a beaker of 70% alcohol to sterilise them. Always replace these into the alcohol in between manipulations later on.
Immerse the leaf in 1% chlorine (bleach) solution in a sterile plastic petri dish for 15 minutes. Keep the lid on the petri dish after each manipulation of the leaf from now on.
Pour off the solution. Cover the leaf with about 20 ml of 70% ethanol, replace the lid, and swirl gently for about 1 minute.
Pour off the solution. Cover the leaf with sterile water, replace the lid, and swirl gently for about one minute. Repeat this rinsing step three more times. Remember to replace the lid between washings!
Pour off the last rinse water and sterilise the scalpel blade by removing from the alcohol and passing through a bunsen flame. The alcohol will burn off. Allow the blade to cool for a few seconds.
Cut the leaf into 10 mm square pieces. and put the scalpel back in the beaker of alcohol.
Flame the tweezers, allow to cool for a few seconds and transfer the squares onto sterilised culture medium in a petri dish
Incubate at 25oC with 16 hours light / 8 hours dark. Use artificial light on a timer or just place somewhere warm in the classroom.

Records: Keep a diary. Note any contamination. How well did your tissue culture experiment work?

70% ethanol solution is made by mixing 70 ml ethanol with 30 ml water.
Sterile water can be prepared in a pressure cooker (15 minutes at 15 lbs pressure)
Pre-sterilised plastic petri dishes are available from most biomedical supply companies. An alternative is to use glass petri dishes with matching lids that can be sterilised by baking in an oven at 250oC for 1 hour.
Culture medium available in powder form from Sigma

FIND OUT MORE:-

What other type of plants are commonly grown in tissue culture?
What are the benefits of growing from tissue culture compared to growing from seed?
Tissue culture is important for growing viruses. What are these viruses used for?