Deoxyribonucleic acid or DNA is the genetic material of living beings . The extraction of DNA is done with the purpose of doing genetic, forensic and molecular biology studies. We can extract DNA easily with homemade products and a little care.
For more professional genetic studies, special protocols with different steps and materials that may be contaminants are required. However, we can do a DNA extraction from materials that we have at home, or what we can do in the school’s biology lab.
Objectives of DNA extraction
DNA is the genetic material of living beings.
DNA consists of two chains of nucleotides, joined together forming a double helix. The skeleton has phosphate groups that are negatively charged and polar, which gives the DNA a negative net charge. Within the nucleus, this molecule is packed with proteins around it.
The objectives of this DNA extraction are:
- Apply a simple technique to obtain DNA from plant cells.
- Understand the process necessary for DNA extraction.
The isolation of DNA using the protocol with common salt and dishwasher is a simple, easy, fast, economical and non-polluting alternative. It is based on the physicochemical characteristics of the nucleic acid.
Materials to perform the DNA home extraction
- Plant material, preferably strawberries ( Fragaria vesca ), kiwi ( Actinidia chinensis ) or bananas ( Musa sp .).
- Mortar or blender.
- Scalpel or knife.
- Bath of María (37ºC).
- Gauze or cotton.
- 100 ml test tube or measuring cup.
- Test tube or small bottles.
- Distilled water.
- Dishwashing detergent
- Cooking salt or sodium chloride NaCl.
- Filter paper and strainer.
- Ethanol or isopropyl alcohol with a concentration higher than 70%, cold (5ºC).
The procedure to perform DNA isolation follows the following steps:
Step 1: preparation of saline-soap solution
Prepare a saline-soap solution composed of 100 ml of water (half a cup), 10 ml of dishwashing detergent (1 tablespoon) or 20 ml of shampoo (without conditioner, two tablespoons) and 13 g of NaCl or kitchen salt (one tablespoon) ).
Step 2: preparing the fruits
If we use kiwi or banana, we peel and cut it into small cubes, then grind it in a mortar or liquefy it. If we use strawberries, we can place them in a bag with airtight seal and crush them.
Step 3: adding the salt and detergent solution to the fruit
In the mortar with the crushed fruit (or in the bag with the strawberries) pour the saline-soapy solution and continue the process of breaking the fruits. Optionally, we can place the preparation in a water bath at 37ºC for 15 minutes to facilitate the extraction
Step 4: separation of solid material, proteins and lipids
The salty-soapy fruit juice is strained and filtered to remove the solids, proteins and lipids from the aqueous solution of the DNA.
We passed through a strainer or filter paper to remove the thickness of the fruit soup and then, through the gauze or cotton, to obtain 5 ml of the filtered liquid in a test tube or 50 ml in a jar.
Step 5: precipitation of DNA by alcohol
Between the aqueous phase and the alcohol phase begins to observe the DNA fibers.
On the fruit juice, pour slowly through the walls of the ethanol or cold isopropanol test tube. Let rest a few minutes. A whitish, gelatinous layer should appear.
If we introduce a glass rod or a wooden stick, with circular motions we can recover the DNA wrapped in the rod.
Explanation of the stages of DNA extraction
When we conduct an experiment in science, it is always essential to understand the basis of the techniques and why the phenomena occur. Let’s see the explanation of each stage.
Choice of plant material
Plants have three types of DNA: nuclear, mitochondrial and chloroplast. In addition, most plants have polyploidy , that is, the condition of some cells or organisms possessing more than one set of chromosomes.
Mature strawberries (Fragaria x ananassa) are an excellent material to extract DNA. They are easy to crush and contain enzymes, called pectinases and cellulases, that help break down cell walls. In addition, strawberries are octoploid (7 chromosomes x 8 = 56 chromosomes), that is, they have eight copies of each chromosome, which provides more DNA than banana or kiwi.
The kiwi has 29 chromosomes and is tetraploid. The banana or banana has 11 chromosomes and the majority is triploid. But you can also do this experiment with onions, lentils or other vegetable.
Rupture of tissues, walls and cell membranes
The detergent molecules help break down the cell membranes and release the DNA from the nucleus.
By friction with the mortar or by liquefying the material, the junctions between the cells and the cell wall are broken. This homogenization facilitates the lysis or rupture effect that helps to release the genetic material.
Soap or liquid detergent helps break down cell membranes, which are composed of lipids.
Elimination of proteins and lipids
Non-soluble components such as fibrous material and proteins are separated from DNA by filtration.
After the lipids and proteins are removed, the phosphate groups in the DNA are negatively charged and polar. DNA dissolves in aqueous solutions, but it is insoluble in alcohol.
The addition of ethanol and high concentrations of sodium ions that bind to the phosphate groups, reduces the repulsive forces between the chains and allows the DNA to fold on itself making it insoluble. In this way it precipitates and forms a whitish filamentous mesh containing proteins and other materials.
Break the cell walls and separate the cells.
What is the role of the detergent addition?
The dishwasher or shampoo helps dissolve the cell membrane that is a lipid bilayer.
What is the role of sodium chloride?
Sodium and chloride ions neutralize negative DNA charges.
Sodium chloride help to separate some of the proteins that are attached to DNA. It also keeps dissolved proteins in the aqueous layer to prevent them from also precipitating with alcohol, along with DNA. Additionally, the sodium (Na + ) cations counteract the negative charges of the DNA phosphates.
What is the role of alcohol?
Alcohol decreases the dielectric constant of the aqueous solution. The cold ethanol causes the DNA to separate from the water and precipitate. When the DNA separates from the aqueous solution, it tends to clump together, which makes it visible. The long strands of DNA will wrap around the rod as the interface between the two layers is scrambled.