P12. DNA EXTRACTION

INTRODUCTION:

• Desoxyribonucleic acid (DNA)  is a nucleic acid that encodes the genetic instructions used in the development and functioning of all known living organisms and many viruses.

• Nucleic acids are biopolymers formed by simple units called nucleotides. Each nucleotide is composed of a nitrogen-containing nuclease (G, T, C, A) as well as a monosaccharide (desoxyribose) and a phosphate group.

• These nucleotides are joined to one another in a chain by covalent bonds between the sugar of the nucleotide and the phosphate of the next.

• Most DNA molecules consist of two strands coiled around each other to form a double helix. The two strands run in opposite directions to each other and are therefore anti-parallel. Moreover the bases of the two opposite strands unit according to base pairing rules : A-T and G-C.

•  Within cells, DNA is organized into structures called chromosomes.

OBJECTIVES:

1. Study DNA structure
2. Understand the process of extracting DNA from a tissue.

MATERIALS:- 1L Erlenmeyer flask.- 100mL beaker.- 10mL graduated cylinder.- Small funnel.- Glass stirring rod.- 10mL pipet.- Knife.- Safety goggles.- Cheesecloth.- Kiwi.- Pineapple juice (1mL/5mL).- Distilled water.- 90% Ethanol ice-cold.- 7mL DNA buffer.- 50mL dish soap.- 15g NaCl.- 900mL tap water.

PROCEDURE:

Put the ethanol in the freezer, you will need it really cold later!Prepare the buffer in 0,5L beaker: Add 450mL of tap water, 25mL of dish soap and 7g NaCl. Stir the mixture.

1. Peel the kiwi and chop it to small pieces. Place the pieces of the kiwi in one 600mL beaker and smash with a fork until it becomes a juice pure.2. Add 8mL of buffer to the mortar. 3. Mash the kiwi puree carefully for 1 minute without creating many bubbles. 4. Filter the mixture: put the funnel on top of the graduated cylinder. Place the cheesecloth on top of the funnel.5. Add beaker contain carefully on top of cheesecloth to fill the graduated cylinder. The juice will drain through the cheesecloth but the chucks of kiwi will not pass through in to the graduated cylinder.6. Add the pineapple juice to the green juice ( you will need about 1mL of pineapple juice to 5mL of the green mixture DNA solution). This step will help us to obtain a purer solution of DNA . Pineapple juice contains an enzyme that breaks down the proteins.7. Tilt the graduated cylinder and pour in an equal amount of ethanol with an automatic pipet. Put the ethanol through the sides of the graduated cylinder very carefully. You will need about equal volumes of DNA solution to ethanol. 8. Place the graduated cylinder so that it is eye level. Using the stirring rod, collect DNA at the boundary of the ethanol and kiwi juice; only the stir in the above ethanol layer!9. The DNA precipitate looks like long, white and thin fibers.10. Gently remove the stirring rod and examine what the DNA looks like. 

QUESTIONS:1.- What did the DNA looks like?

• The DNA looks like white  and thin fibers.

2.- Why do you mash the kiwi? Where it's located inside the cells?

• The DNA is located in the nucleus and we mashed it to liberate it. 

3.- Explain what is the function of every compound of the buffer (soap and salt). 

• The salt can breaks the nucleus cell and we put soap to take away the proteins.

4.- DNA is soluble in water, but not in ethanol. What does this fact have to do with our method of extraction?

• We can see the DNA in the part of ethanol because, if we touch the water the DNA can dissolve.