EXPERIMENT No.

12
NUCLEIC ACIDS

OBJECTIVES: At the end of this experiment, students are expected to learn how to:
1. Extract nucleic acid from liver.
2. Perform Qualitative test to detect components of nucleic acids.

ASSIGNMENT
A. Complete the table below with expected results for positive test of the following
substances.

SUBSTANCES TEST/REAGENTS INDICATION OF POSTIVE

RESULT
Purine

Pentose

Deoxyribos

Phosphorous

B. Draw that structure of DNA and RNA then label the components as sugar, base, and
phosphate.

DNA RNA
DISCUSSION
Biological systems employ large molecules, the nucleic acids, for the essential functions
of storage and transmission of genetic information. With the exception of certain viruses, it is
deoxyribonucleic acid (DNA) that serves as the primary genetic material, the gene. Various
DNA and its translocation to primary protein structure.

REAGENTS
Liver 5% Trichloroacetic Acid Ethyl alcohol
Ether 2N HCL 2N NaOH
Acetate buffer 10% copper sulfate Saturated Sodium bisulfate
Orcinol reagent Diphenylamine reagent 1N H2SO4
Concentrated HNO3 Molybdate-napthol sulfonic
acid

APPRATUS: Blender, Graduated cylinder, Centrifuge, Test Tubes, Glass rod, Beaker, Test
Tube holder, Test Tube rack, Hot Plate, Water Bath, Tong, Thermometer, Evaporating dish,
Dropper, Pasteur pipette.
PROCEDURE
A. EXTRACTION OF NUCLEIC ACIDS FROM LIVER
Liver is suspended in 5% Trichloroacetic acid (TCA) to extract the acid soluble substances.
The residue is extracted with alcohol-ether mixture to remove the lipids. From the residue
DNA and RNA extracted with hot 5% TCA
1. Homogenized the liver with a volume of water.
2. Place 1mL of the homogenous mixture in the test tube.
3. Add about 5mL of 5% TCA, stir and centrifuge for 5 minutes.
4. Decant and discard the supernatant liquid. Repeat the washing using alcohol-ether
mixture 3:1.
5. Suspend the residue in 7mL of 5% TCA and heat at 90C for 15min with occasional
stirring.
6. Cool, centrifuge and decant the supernatant to a clean test tube and add water to make
up to 10 mL. Use this solution to test for purine and determination of RNA and DNA.

B. TEST FOR PURINE

1. Place 1mL of the solution prepared in Part A in a test tube and add 1mL of 2N HCl.
2. Prepare a similar tube containing 1mL of water and 1mL of 2N HCl (Blank).
 3. Place both tubes in a boiling water bath for 20mins. This hydrolyzes the nucleic acid to
form free purines.
4. Add 1mL of 2N NaOH and 2mL acetate buffer to the tubes.
5. Again place in boiling water bath and heat
6. Add 0.5mL of 10% copper sulfate. A bluish-brown precipitate forms in both tubes.
7. Add 0.5mL of saturated sodium bisulfate and mix.
8. Heat in a water bath for a few minutes. Observe.
A white or light tan flocculent should be present in the tube containing the purine solution.
C. DETERMINATION OF RNA- ORCINOL TEST FOR PENTOSES

1. Dilute 0.5mL of the solution prepared in part A with 2.5mL of water.

2. Add 3mL of the Orcinol reagent and heat in the water bath.
3. Cool the tube under tap water and note the result.

D. TEST FOR PHOSPHOROUS

Precaution: To avoid irritating vapors do this procedure under the hood or near the window.
1. Evaporate 0.5mL of the prepared solution in part A mixed with 1mL of H2SO4 until
the mixture chars or blackens.
2. Cool and add 2 drops of concentrated HNO3.
3. Heat until brown fumes disappear and the solution clears.
4. If the solution does not clear, allow to cool and add 2 drops of concentrated HNO3 and
heat.
5. To the residue, add water to make 2mL volume.
6. Add 1mL of the molybdate-napthol sulfonic acid reagent.
7. Note the result.
DATA/RESULTS
TEST OBSERVATION
A. Test for Purines
Note the color of precipitate

B. Orcinol Test for Pentoses in RNA

C. Diphenyl Test for Deoxyribose in

DNA

D. Test for Phosphorus

CONCLUSION:
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POST-LABORATORY QUESTIONS: USE THE BACK PAGE IF NECESARRY

1. What is sickle cell disease? What causes it? What is the difference in the shapes of the
red blood cells? What effect does this have on their ability to carry oxygen?
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2. The insertion of wrong nucleotide into a DNA code group could result in? Maturation,
recombination, splicing or cloning? Explain.
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