Class 12 Biology - Molecular Basis of Inheritance

Activity 5.1: Isolation of DNA from Plant Material (Onion/Banana)

Aim: To extract and visualize genomic DNA from a plant source using common laboratory reagents.

Materials Required:

50g fresh Onion bulbs or ripe Banana
Lysis Buffer: 10 mL Liquid Detergent (SDS-based), 3g Sodium Chloride ( $NaCl$ ), and 90 mL Distilled water.
Chilled 95% Ethanol or Isopropanol (stored at -20°C for at least 2 hours)
Meat tenderizer (as a source of Protease) or 5% Pineapple juice
Mortar and pestle
Cheesecloth or fine muslin cloth
Glass rod or spooling loop
Beakers (100 mL and 250 mL)
Water bath (maintained at 60°C) and Ice bath

Procedure:

Mechanical Lysis: Macerate 50g of onion in a mortar and pestle with 20 mL of lysis buffer until a fine slurry is formed.
Thermal Incubation: Transfer the slurry to a beaker and heat in a water bath at 60°C for exactly 15 minutes.
Cooling: Immediately transfer the beaker to an ice bath for 5 minutes to prevent DNA degradation.
Filtration: Filter the mixture through a cheesecloth into a clean beaker. Collect the filtrate (containing DNA, proteins, and RNA).
Proteolysis: Add 5 mL of 5% Pineapple juice (or a pinch of meat tenderizer) to the filtrate. Mix gently and let it sit for 5 minutes at room temperature.
Precipitation: Carefully pour 20 mL of ice-cold 95% ethanol down the side of the beaker so it forms a distinct layer on top of the filtrate. Do NOT mix.
Spooling: Wait for 2-3 minutes. White, thread-like fibers of DNA will precipitate into the ethanol layer. Use a glass rod to "spool" the DNA by rotating it gently at the interface.

Scientific Note The detergent (SDS) disrupts the phospholipid bilayers of the cell and nuclear membranes. High salt concentration ( $NaCl$ ) helps neutralize the negatively charged phosphate backbone of DNA, allowing it to aggregate. Ethanol has a lower dielectric constant than water; in its presence, the electrostatic attraction between $Na^{+}$ and $PO_4^{3-}$ increases, causing the DNA to precipitate out of the aqueous solution.

Safety First Ethanol is highly flammable. Ensure all heating elements (hot plates/water baths) are turned off before bringing chilled ethanol near the experimental bench.

Observation: A white, cloudy, viscous mass of fibers appears at the interface of the two liquids, which can be wound around a glass rod.

Lab Best Practice The 15-minute incubation at 60°C is critical. It denatures DNase enzymes that would otherwise chop up the DNA. However, exceeding 15 minutes or higher temperatures will start denaturing the DNA itself.

Activity 5.2: Visualizing the Double Helix (Model Building)

Aim: To construct a scale model of the B-DNA double helix to understand its physical dimensions.

Materials Required:

Colored beads (4 colors: A-Red, T-Green, G-Blue, C-Yellow)
Flexible wire or plastic tubing
Spacers (measuring 0.34 nm and 2.0 nm in scale)
Vernier calipers

Procedure:

Maintain a constant scale (e.g., 1 cm = 0.1 nm).
Space the nitrogenous base beads exactly 3.4 cm (representing 0.34 nm) apart along the wire.
Ensure each full turn of the helix contains 10 base pairs (34 cm per turn).
Maintain a helix diameter of 20 cm (representing 2.0 nm).

Scientific Note The B-form of DNA, described by Watson and Crick, is a right-handed helix. The stability of the helix is maintained by hydrogen bonds between complementary bases (2 for A=T, 3 for G $\equiv$ C) and base-stacking interactions (van der Waals and hydrophobic forces) between the planar aromatic rings of the bases.

Activity 5.1: Isolation of DNA from Plant Material (Onion/Banana)

Aim: To extract and visualize genomic DNA from a plant source using common laboratory reagents.

Materials Required:

50g fresh Onion bulbs or ripe Banana

Lysis Buffer: 10 mL Liquid Detergent (SDS-based), 3g Sodium Chloride (

NaCl

), and 90 mL Distilled water.

Chilled 95% Ethanol or Isopropanol (stored at -20°C for at least 2 hours)

Meat tenderizer (as a source of Protease) or 5% Pineapple juice

Mortar and pestle

Cheesecloth or fine muslin cloth

Glass rod or spooling loop

Beakers (100 mL and 250 mL)

Water bath (maintained at 60°C) and Ice bath

Procedure:

Mechanical Lysis: Macerate 50g of onion in a mortar and pestle with 20 mL of lysis buffer until a fine slurry is formed.

Thermal Incubation: Transfer the slurry to a beaker and heat in a water bath at 60°C for exactly 15 minutes.

Cooling: Immediately transfer the beaker to an ice bath for 5 minutes to prevent DNA degradation.

Filtration: Filter the mixture through a cheesecloth into a clean beaker. Collect the filtrate (containing DNA, proteins, and RNA).

Proteolysis: Add 5 mL of 5% Pineapple juice (or a pinch of meat tenderizer) to the filtrate. Mix gently and let it sit for 5 minutes at room temperature.

Precipitation: Carefully pour 20 mL of ice-cold 95% ethanol down the side of the beaker so it forms a distinct layer on top of the filtrate. Do NOT mix.

Spooling: Wait for 2-3 minutes. White, thread-like fibers of DNA will precipitate into the ethanol layer. Use a glass rod to "spool" the DNA by rotating it gently at the interface.

Safety First Ethanol is highly flammable. Ensure all heating elements (hot plates/water baths) are turned off before bringing chilled ethanol near the experimental bench.

Observation: A white, cloudy, viscous mass of fibers appears at the interface of the two liquids, which can be wound around a glass rod.

Activity 5.2: Visualizing the Double Helix (Model Building)

Aim: To construct a scale model of the B-DNA double helix to understand its physical dimensions.

Materials Required:

Colored beads (4 colors: A-Red, T-Green, G-Blue, C-Yellow)

Flexible wire or plastic tubing

Spacers (measuring 0.34 nm and 2.0 nm in scale)

Vernier calipers

Procedure:

Maintain a constant scale (e.g., 1 cm = 0.1 nm).

Space the nitrogenous base beads exactly 3.4 cm (representing 0.34 nm) apart along the wire.

Ensure each full turn of the helix contains 10 base pairs (34 cm per turn).

Maintain a helix diameter of 20 cm (representing 2.0 nm).

Class 12 Biology - Molecular Basis of Inheritance (Activities)

Class 12 Biology - Molecular Basis of Inheritance

Activity 5.1: Isolation of DNA from Plant Material (Onion/Banana)

Activity 5.2: Visualizing the Double Helix (Model Building)

On this page

Class 12 Biology - Molecular Basis of Inheritance (Activities)

Class 12 Biology - Molecular Basis of Inheritance

Activity 5.1: Isolation of DNA from Plant Material (Onion/Banana)

Activity 5.2: Visualizing the Double Helix (Model Building)

On this page