The purpose of the experiment was to experience firsthand the isolation of DNA form a plant tissue without destroying its structure and sequence. A white onion was used for the experiment. After several processes, DNA isolate was the visible result. Different chemical tests were performed on the DNA isolate, namely: Dische Test, Murexide Test, Wheeler-Johnson test and Test for Phosphate. Visible results were then noted.
DNA (deoxyribonucleic acid) was discovered in the late 1800s, but its role as the material of heredity was not elucidated for fifty years after that. It occupies a central and critical role in the cell as the genetic information in which all the information required to duplicate and maintain the organism.
Today, Scientists can analyze DNA from minute samples of blood, hair roots and saliva, but before they can analyze DNA, scientists must be able to extract it. The process of DNA extraction is the first step for many laboratory procedures. Scientists must be able to isolate the DNA from the other unwanted substances of the cell gently enough so that the DNA is not broken up.
For this experiment, an onion was used because of its low starch content, which allows the DNA to be more clearly seen. The salt shields the negative phosphate ends of DNA, which allows these ends to come closer so that the DNA can precipitate out of a cold alcohol solution. The homogenizing solution causes the cell membrane to break down by dissolving the lipids and proteins of the cell, which disturbs the bonds that hold the cell membrane together. The homogenizing solution then forms complexes with the lipids and proteins causing them to precipitate out of the solution.
After the isolation of the DNA from the solution, different chemical tests were then performed on the isolate, which were: The Dische Test, Murexide Test, Wheeler-Johnson Test and Test for Phosphate.
The Dische test is a test done to identify DNA. It is the reaction between Dische reagent & 2-deoxypentose yields a blue colored solution.
The Murexide test a test in which the treatment of a substance, usually urine, with nitric acid and ammonia indicates the presence of uric acid by formation of murexide. Positive results form this test yields Purines form Yellow to Red-Violet.
The Wheeler-Johnson Test is a qualitative test for the pyrimidine bases cytosine and uracil, which produces a green coloration when the sample is treated with bromine water. The addition of barium hydroxide will turn the liquid purple.
The Test for phosphate is used to detect phosphate in the DNA, positive reaction yields yellow precipitate.
RESULTS AND DISCUSSION
The following procedures were done in order to isolate the DNA from the onion. First, the onions were minced and then weighed to 25g. The onions were minced because by mincing the onions, it increase its surface area that helps make the membrane at the surface easier to dissolve, and also allows for more efficient absorption of heat and solutions.
Next it was added to a 50ml homogenizing solution, this was done because the homogenizing medium, made of 5% SDS, 0.15M NaCl, 0.15 M sodium citrate and 0.001 M EDTA, was similar to a detergent. It helped break up the phospholipid bilayer of the cells' plasma membranes and nuclear envelopes. When in SDS, the proteins and lipids of the membrane, but especially the lipids, are broken down because the SDS causes the bonds holding the membrane together to break. The lipids, which are repelled by the SDS, separate and break up the membrane. This is why detergent is used to remove a grease stain; grease is a lipid. The EDTA helped weaken the membrane as well, making the DNA that was in the nucleus now available for extracting.
Then the solution together with the onion was heated at 60 degrees, this method was used to speed the breakdown process by energizing the molecules, and also, hopefully help to...