Experiment 7: To Study Pollen Germination on Stigma Through a Permanent Slide

AIM

To study the pollen germination on stigma through permanent slide.

PRINCIPLE

Pollen germination on stigma is an essential step of sexual reproduction in angiosperms. Compatible pollen grains are recognized by stigma and germinate to form pollen tube that carries male gametes toward ovule.

Mature pollen grain contains vegetative cell and generative cell enclosed within exine and intine. During maturation, microspore undergoes asymmetric division forming large vegetative cell and small generative cell with dense cytoplasm.

On compatible stigma, intine emerges as pollen tube through germ pore. Tube nucleus guides growth and generative cell divides to form two male gametes. One fuses with egg cell and other fuses with secondary nucleus.

REQUIREMENTS

• Permanent slide of pollen germination with stigma
• Compound microscope
• Drawing sheets
• Pencil and eraser
• 5-6 excised styles with stigma of Petunia / grass / maize / sunflower for temporary slide preparation
• Beaker
• Water
• Slides
• Cover slips
• Cotton blue stain
• Brush and needle
• Glycerine

PROCEDURE

Using Temporary Slide:

1. Place the freshly excised stigma and styles in boiling water in a beaker for 5-10 minutes. This will soften the tissues and also prevent them from drying.
2. Add cotton blue dye using a dropper onto the tissue placed on a glass slide.
3. Wait for 3-5 minutes and then wash-off the excess stain with water.
4. Mount one stigma in a drop of glycerine on a slide.
5. Place a cover slip on the stigma and gently press the cover slip on the material.
6. Place the temporary slide under a compound microscope and observe initially under low magnification followed by higher magnification.
7. Draw a well-labelled diagram of the structures seen.

Using Permanent Slide:

1. Pick up a good permanent slide of pollen germination.
2. Place it under low power of microscope and observe the slide.
3. Focus structures seen in low power, then in high power to have clear view of details of pollen tube.
4. Draw a well-labelled diagram of the structures seen.

OBSERVATION

The slide shows the presence of many germinating pollen grains on the stigma of the carpel. Pollen grains adhere to stigma surface and show tube emergence.

The pollen tubes of the germinating pollen grow through the style, traversing the tissues of the style. The slide shows many such growing pollen tubes of varied length.

Key structures identified in the slide:

• Pollen grain tetrad formed after meiosis of microspore mother cell.
• Stages of microspore maturation showing vacuoles, nucleus and asymmetric spindle formation.
• Mature pollen grain with exine, intine, vegetative cell and generative cell.
• Vegetative nucleus, generative nucleus, tube cell, tube nucleus and germ pore clearly visible.
• Pollen tubes penetrating stigmatic tissue and growing through style toward ovule.

Additional Illustrative Observations for Different Species:

RESULT

As the pollen tube grows into style, it reaches to the ovule and carries the two male gametes; one to the egg cell and another to secondary nucleus for fertilisation to occur inside the ovule.

CONCLUSION

Pollen germination on compatible stigma is essential for double fertilisation in angiosperms. The study confirms that pollen tube carries male gametes through style to ovule, ensuring syngamy and triple fusion characteristic of flowering plants.

PRECAUTIONS

• The prepared permanent slide should be handled carefully.
• Draw proper figures of the structures as seen under low and higher magnifications of the compound microscope.
• Label the figures accurately.