Pineapple Smoothie DNA Isolation Laboratory Report

Pineapple Smoothie: DNA Isolation

Bergil Bernaldo, Bos Mauro Brandis, Jim Pauline Guiyab, Krizelle Ann Hugo, Sheryll Ann Trinidad

Abstract  Deoxyribonucleic acid is the genetic material found in all cellular organisms. To find out  how the DNA looks like macroscopically, DNA isolation was performed on a pineapple fruit. The isolation starts with breaking down of the fruit by turning it into smoothie, next is the addition of   ethanol in the pineapple puree destroys the lipids in the cells and separating the DNA. The experiment had a control group using -naphtol, ammonium molybdate and phosphoric acid for  the positive control. The presence of DNA was determined by the change in color turning from white white to deep deep blue blue due due to the the reac reacti tion on of the the ammo ammoni nium um moly molybd bdat atee and -nap -napht htol ol with with the the  phosphates acting as the backbone of the DNA. The experiment yielded a positive result  confirming the presence of DNA in the isolated spooled material.

Keywords Deoxyribonucleic acid; Pineapple ( Anasas comosus); nucleoproteins; comosus); nucleoproteins; -naphtol; Ammonium molybdate;

Introduction All known cellular life and some viruses contain DNA  DNA  (deoxyribonucleic acid). acid). DNA is a nucleic acid that contains the genetic instructions for the development and function of living things. [1] It has a double helix structure and is made up of chemical building blocks called nucleotides. These building blocks are made of three parts: a phosphate group, a sugar group and one of four types of nitrogen bases. [2] The four types of nitrogenous bases are; adenine (A), guanine (G), thymine (T), and cytosine (C). The Biochemistry Biochemistry students of BS Chemistry Chemistry 3-1 were asked to isolate isolate DNA from pineapple pineapple (Ananas comosus) fruit. comosus) fruit. A pineapple fruit is actually multiple fruits that have been formed from a thickened stem surrounded by hundreds of fruitlets that have been fused together. Pineapples have a tough and hard rind, which consists of the remains of the flowers and the pointed bracts. The color is golden orange and, despite its hard and dry appearance, juice can be extracted from the rind. Pineapple has no seeds but the core of the fruit is hard since it consists of the old stem. Removal of the core produces the characteristic pineapple rings. The flesh is juicy and yellow.  [ 3] 1

DNA isolation is the process of extracting DNA from a cell. It involves separating DNA from other cellular components. DNA is isolated by placing the cells in a tube containing a special solution and mechanically or chemically breaking the cells open. This causes the cell to release its contents into the solution which may contain enzymes, chemicals, and salts. Enzymes are used to chew up the proteins; chemicals to destroy any RNA present and salts to help pull the DNA out of solution. The DNA will exist in long strands that form a mucous-like glob.[4] A phosphate test using ammonium molybdate and α-naphthol can be used to confirm the  presence of DNA.

Methods START

Collect the ethanol that contains the precipitate and place in a separate test tube.

Prepare chunks of pineapple then  puree.

Filter the puree by using cheesecloth then collect the filtrate

Run the sample in the centrifuge for 2 minutes.

Dispose the ethanol to isolate the s ooled material.

Add liquid detergent to destroy the lipids.

Add 3-5 drops of Sulfuric Acid.

Put the sample in a test tube. It should occupy 1/3 of the test tube.

Add 3-5 drops of Nitric Acid then a l constant heat.

Add ice cold 95% Ethanol.

 No Submerge the sample in a water   pan that contains ice.

Does the color of the sample become clear pale yellow?

Let the sample sit for a couple of  minutes.

Yes Add distilled water and some drops of  Ammonium Molybdate. Do you observe white precipitate in the ethanol?

 No

Does the color of the sample become clear?

2

Yes

 No

A

Add α-napthol

Apply heat until α-napthol dissolves.

Compare the result with the control set-up.

B C

End

Start Start

Put 1ml of Phosphoric Acid in a test tube.

Put 1ml of distilled water in a test tube.

Add 1ml of Ammonium Molybdate and a small piece of  α-napthol.

Add 1ml of Ammonium Molybdate and a small piece of α-napthol.

Heat until α-napthol dissolves. Heat until α-napthol dissolves.

Add water to dilute B C

3

Results and Discussions A.

Liberation of Nucleoproteins For approximately two minutes, white spool-fiber known as “nucleoproteins” precipitated in the ethanol layer of the test tube. The nucleoproteins contain the DNA material needed in the following test.

B. Confirmation for presence of DNA The clear color of the nucleoprotein aliquot remained clear after few drops of sulfuric acid was added. Upon heating the test tube, drops of nitric acid was added until yellow fumes disappeared turning the color of the solution into light yellow. The color of the solution turned clear upon heating it with few drops of ammonium molybdate. The solution turned dark blue when heated after a small amount of α-naphtol was added. The solution was diluted to visualize better the blue color that indicates a positive result. The presence of the DNA is confirmed because of the phosphate backbone in the DNA that reacted with the reagents.

Figure 1: Blue color of the positive Figurecontrol 2: Addition of nitric acid in acid digestion. (left) and blue color of the positive result (right).

References [1] DNA. http://www.sciencedaily.com/articles/d/dna.htm (accessed December 7, 2013) [2] Deoxyribonucleic Acid (DNA) Fact Sheet. http://www.genome.gov/25520880 (accessed December 7, 2013) [3] Characteristics of a Pineapple | eHow. http://www.ehow.com/info_8553867_characteristics pineapple.html (accessed December 8, 2013) [4] http://oklahoma4h.okstate.edu/aitc/lessons/intermed/DNA_Blueprint.pdf  4