A Comparison of Different Tissue Culture Techniques Used to Produce Dionaea muciapala, the Venus Fly Trap.


Hypothesis
Background
Procedure
Growing Info.
Data
Conclusion
Bibliography

Purpose

The purpose of this study was to determine which tissue culture technique (root, leaf, stem) produces the tallest plantlet in a given time.


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Hypotheses

Hypothesis 1: Leaf culture technique would produce the tallest plantlet in a given time.
Hypothesis 2: Root cultures would produce the smallest plantlet.
Hypothesis 3: Stem cultures would have the highest contamination rate.

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Background

Venus fly traps, scientific name Dionaea muciapala are a type of carnivorous plant, this means they are capable of digesting and absorbing nutrients from organic matter. Basically they eat insects such as mosquitoes, flies, beetles, and any other insect that crawls or flies into its trap. Venus fly traps catch there food by attracting the insects with a sweet fragrance, the plant also has red in the middle of the trap so it is very noticeable. When the insect flies into the trap and touches two sensitive hair or one sensitive hair twice the trap closes and traps the insect. This closing is caused by the tension in the traps being released and closing on the insect. Then the digestive enzymes go to work digesting the insect, and then the cell wall on the inside of the trap absorbs the digested material. The plant needs the insects because these carnivorous plants live in acid bogs which are bogs with sandy soil and very little minerals in the soil. Insects are high in nitrogen, and many important minerals for the plant. The Venus fly traps use the minerals from the insects to survive. In spring the plants produce greenish-white flowers, and in winter some may become dormant. There are many different clones of Venus fly traps. There are types that turn completely red when in full sun, types that are all green, types with very long "teeth", and others that have short stubby "teeth". There is only one species of Venus fly trap, so there are not any hybrids, there are only clones.

More than 4,000,000 Venus fly traps are cultivated each year for sale by major wholesalers and retailers. The green swamp preserve in Brunswick county, North Carolina use to have 200,000 acres of Venus fly trap habitat but now there is only 16,000 acres of Venus fly trap habitat. People are always trying to find newer and easier ways to culture Venus fly traps so that they don't have to harvest them from the wild..

Tissue culture is a way to vegetative propagate plants using small cuttings to produce more than one plant; some times thousands. Some of the most common ways to culture plants are root propagation, leaf propagation, bulb propagation, seed propagation, and steam propagation. Tissue culturing is a relatively new way to propagate plants. Many large plant companies use tissue culture. The cultures are usually relatively hardier, healthier, and sometimes even cheaper than plants produced by other methods. Some people prefer cultured plants. Some plant saving projects use tissue culture to replace some of the plants in the wild. The best part about tissue culture is that if you use multiplication media the plants will keep multiplying until you stop them and then transfer them into pretransplant media, which stops multiplication and starts root development.

Venus fly traps are not the only carnivorous plants, there are Sundews (Drosera) and Butterworts (Pinguicula) which use a sticky substance to capture insects. Pitcher plants (Sarracenia) which use a pitfall trap and Blatterworts (Utricularia) which use a mechanical tra, and are sometimes found in water.

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Procedure


Sterilize
1.) Sterilize beakers, and scalpels in a baby food jar sterilizer.
2.) Sterilize tweezers in a graduated cylinder of ethanol.
3.) Sterilize tissue by placing the 1 inch tissue fragments in a culture dish with about 20 ml of ethanol for 30 sec., then place fragments in a culture dish with about 20 ml of a 20% solution of bleach, then do 3-5 min. washes in sterile water.

2. Prepare Work Area
1.) Place culture tubes rack with tubes in the flow chamber.
2.) Fill a 200 ml and 100 ml beaker with ethanol then place in the flow chamber.
3.) Fill a 200 ml and 100 ml beaker with sterile water then place in the flow chamber.
4.) Place the scalpel in the 100 ml beaker of ethanol.
5.) Place the tweezers in the 200 ml beaker of ethanol.

7.) Spray down inside of chamber with isopropyl alcohol.

3. Culture
1.) Dip the scalpel in the 100 ml beaker of water, and dip the tweezers in the 200 ml beaker of water.
2.) Cut off any brown edges on the tissue.
3.) Dip the tweezers in the ethanol and into the water.
4.) Place one tissue fragment into each culture tube.
Repeat the procedure for each type of tissue.

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Growing Venus Fly Traps

1. Keep in the culture tubes until the the Venus Fly traps are 5 cm tall.
2. Then transplant to potting media(a good media for Venus Fly Traps is milled sphagmum moss).
3. To keep the humidity high a good technique is to put popsicle sticks on each side and wrap in clear plastic wrap. The popsicle sticks keep the plastic wrap off the growing plants.
4. Never allow the soil to dry out.
5. Keep in full sun.
6. Do not fertilize, because fertilize can cause serious damage to the plant and maybe even kill it.

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Data

Four runs were conducted, the preliminary results were trial 1: 100% contamination for leaf cultures, 100% contamination for the stem cultures, and 0% contamination for root cultures. Trial 2: 70% leaf cultures, 0% stem cultures, 0% root cultures. Trial 3: 70% leaf cultures, 10% stem cultures, and 0 % root cultures. Growth data is not available due to the slow growth rate of the cuttings.




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Conclusion

Due to slow growth rates there was not enough data to accept or reject hypotheses 1 and hypothesis 2.
The data did not support hypothesis 3 and the hypothesis was rejected. Stem cultures did not have the highest contamination rate

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Bibliography

Alexander, Peter et al ,1989, Biology the Living World, Englewood Cliffs, Prentice-Hall Inc.

Arditti, Joseph, 1977, Orchid Biology Reviews and perspectives, 1st Edition, London, Cornell University press.

D Amato, peter, 1998, The Savage Garden, Berkley, Ten speed press.

Schwartz, Randall, 1974, Carnivorous Plants, New York,Avon Books.

Silverstein, A.,et al 1996, The Kingdoms Plants, Brookfiled, Twenty-first Century Books.

Unknown. Carnivorous plants.[Online] Available at http://www.pitcherplant.com, Accessed on Nov. 1,2000

Unknown. Tissue Culture.. [Online] Available at http://www.orchidsource.com, Accessed on Nov. 1, 2000

Unknown. Tissue Culturing.[Online] Available at http://www.avtbiotech.com, Accessed Nov.1 ,2000



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