Practical 01: Study of Plant Cells

Aim: Study of Plant Cells

Cell: The cell (from Latin cella, meaning "small room") is the basic structural, functional, and biological unit of all known living organisms. A cell is the smallest unit of life. Cells are often called the "building blocks of life". The study of cells is called cell biology.

Cells are not only the basic building units of living organisms. They are also the physiological units of a living being. Since there is a close relationship between structure and function it is necessary to understand cell structure in order to understand its physiological functions. In the following experiments, the basic structure of a plant cell as seen under the light and electron microscope will be described.

Several parts of a plant cell can easily be observed under a light microscope, e.g., cell wall, plasma membrane, vacuole, plastids, and nucleus. Due to their relatively smaller size, mitochondria cannot easily be seen with a light microscope but they can be made visible under a light microscope by staining with special chemicals.

Light Microscopy

Materials required:

1. Leaves

2. Aceto-carmine solution (boil 4-5 g carmine in 100 ml of 50% acetic acid and filter after cooling),

3. cone,

4. sulphuric acid,

5. iodine solution (dissolve 0.5-1.0 g of potassium iodide (KI) in 100 ml of water. Then dissolve 1 g of iodine crystals),

6. carmine-alum solution (dissolve l g carmine + 10 g alum (K₂SO₄.Al₂(S0₄)₂.24 H₂O) in 200 ml distilled water by warming.

7. Filter the solution. Add 1 ml of formaldehyde to preserve the stain), neutral-red solution (5 mg in 100 ml water),

8. extremely dilute solution of gentian-violet (the solution should be just lightly colored),

9. Rhodamine B solution (dissolve 0.10 mg rhodamine B in 100 ml of tap water),

10. 2 M sucrose solution

Ordinary compound microscopes and phase contrast microscopes

Procedure:

i) Protoplasm movement:

• Remove few hair cells from the leaves, leaf petioles, or other plant organs

• mount them on a glass slide.

• Put a drop of water and place a coverslip over the material.

• First, observe under the low power of a microscope and select the most suitable cell.

• Observe under the high power the movement of protoplasm.

• Depending upon the plant material, it may be seen that along with the protoplasm also the cell inclusions in the protoplasm are seen moving along with the protoplasm.

ii) Cell wall:

• Cut an onion bulb into four pieces.

• Cut out a small piece from the interior side of the bulb

• peel up carefully an epidermal layer.

• Place the stripe of epidermal cells on a glass slide and mount on a drop of safranin stain.

• Allow stain for about 15 min.

• Wash away excess stain with dilute acids.

• The cell wall stains red.

• In the same way, stain another stripe of the epidermis with a drop of neutral-red solution.

• The middle lamella will be stained red.

iii) Nucleus:

• A Nucleus can distinctly be seen in an epidermal cell of an onion scale as prepared above.

• The Nucleus can be stained by applying a drop of aceto-carmine solution.

• If carefully observed at high power, one can even see nucleoli that are not or only very faintly stained by the stain.

• In addition to epidermal cells of the onion scale, very distinct and big nuclei can also be seen in the hairs (trichome cells) of a cucumber leaf or the epidermal cells of young leaves of Tradescantia.

iv) Plastids:

Chloroplasts:

• Mount thin sections of green leaves of maize, or spinach on a microscopic slide.

• Place a coverslip over the material.

• First, observe under the low power and then under the high power of a microscope.

• In the maize leaf transverse section, one can observe chloroplast dimorphism, i.e., two different kinds of chloroplasts.

• Observe the chloroplasts in the mesophyll cells and the bundle sheath cells.

• Mark the differences between the two types of chloroplasts.

Chromoplasts:

• Chromoplasts are plastids that contain no chlorophyll pigments. They contain yellow (carotenes, xanthophylls) or red (anthocyanins) pigments and are commonly found in fruits (tomato) and flowers.

• Make thin sections of the yellow flower petals (e.g., pansy, roses), red tomato-fruit skins, and carrot.

• Mount them with a drop of water on a glass slide,

• cover with a coverslip and observe carefully, first under low power and then under high power.

• Yellow and red-colored round bodies can be distinctly be seen.

Leucoplasts:

• They are colorless plastids and are found in roots, rhizomes, and different storage organs.

• Peel off lower epidermis from the leaves of Rhoea discolor and mount it in a drop of water on a glass" slide.

• Cover with a coverslip.

• Focus under the high power a distinct nucleus. The nucleus is surrounded by a number of small round bodies, the leucoplasts.

• Replace water on the glass slide with a drop of gentian violet solution for 10-20 seconds. Leucoplasts will appear as violet-colored bodies.

v) Mitochondria:

• Prepare a few stripes of epidermis from onion scales as described above.

• Place them in a bottle containing water.

• Apply suction using an aspirator.

• Take out the stripes,

• place them on a glass slide and treat with few drops of rhodamine-B solution for about 30 min.

• Wash away the excess stain with tap water.

• Observe under a phase-contrast microscope.

• Under green light, mitochondria appear as black rounded structures.

vi) Plasma-membrane and vacuole:

• Plasma-membrane and vacuole can best be observed in epidermal cells containing colored cell sap.

• Peel off an epidermal layer from the leaves

• Discolor and mount it a drop of 2 M sucrose solution.

• Observe under a microscope.

• Due to the concentrated sugar solution, the cells will show the sign of plasmolysis.

• Plasma-membrane becomes distinct and the vacuole is seen full of red pigments, the anthocyanin.