Learning Objective:
To prepare a temporary mount of a leaf peel to show stomata.
Materials Required:
A potted plant of Bryophyllum or Tradescantia
1 Needle
1 Pair of Forceps
1 Watch Glass
1 Dropper
Glass Slides
A (Paint) Brush
Coverslips
1 Blotting Paper
Safranin (red stain)
A pair of scissors/blade
Compound/light/low power microscope
Glycerine
Safety Precautions:
1. HANDLE GLASSWARE WITH CARE AS IF BROKEN, SHARP SHARDS CAN CAUSE INJURIES
2. HANDLE THE NEEDLE WITH CARE
Experimental Accuracy:
1. Ensure leaves being used are freshly plucked
2. Prevent air bubble formation in the slide
3. Place most promising region in the centre of the slide for observation
4. Don’t use too much/little stain
5. Use the lower epidermis, due to a higher concentration of stomata on the underside of the leaf
Methodology:
1) Place a one cell thick layer of cells on the slide?
2) Add a drop of water or stain
3) Lower a coverslip onto the drop(carefully so as to prevent the formation of air bubbles)
4) Place the slide inside folded paper towel/blotting or filer paper to drain excess fluid. Press on the coverslip lightly/gently from the outside of the paper towel
Preparation of Stomata:
Pluck a fresh, healthy leaf from the potted plant. Fold it around the centre(gently) before pulling at it, using a pair of forceps, to obtain a transparent layer – the lower epidermis
Cut the leaf pieces obtained into smaller pieces, so as to allow them to fit onto the microscope slides, using a blade or scissors.
Transfer the pieces to a watch glass containing water. Use a paintbrush to do so(so as to prevent cell damage, while ensuring the epidermis doesn’t dry up).
Piece-by-piece, stain the leaf pieces using Safranin and allow them to remain in the watch glass for a few minutes
Take the pieces out, and place them on clear, glass slides. Add glycerine using a dropper, before gently placing a coverslip on the slide. Place the coverslip using a needle to prevent air bubble formation
Drain excess glycerine/stain using blotting paper
Place the slides under the objective lens, on the stage of the microscope. Use the stage clips to stabilize the slide. Examine the cell.
Observation - Onion Cells:
A row of cells is observed, with features characteristic of plant cells(a nucleus and stomata). The cells that make up this row are epidermal cells. These cells are seen to be tightly packed, with the absence of intercellular spaces. Guard cells can also be observed among the epithelial cells. Gaps between some guard cells are seen(stomata). The cell shape is irregular, however if a different leaf than those specified above is used, then it may be rectangular too.
It can thus be concluded that cells of the lower epidermis, including guard cells, have a nucleus and chloroplast. Guard cells control the opening/closing of stomata. Guard cells are seen to have thicker inner walls, and thinner outer walls.
Experiment in Context:
Plants are adapted in various ways to help them survive. The stomata being more concentrated on the underside of the leaf is one such adaptation. This helps prevent the stomata from interfering with photosynthesis, and losing too much water(excessive transpiration).
Other adaptations include:
Spines: reduce surface area for transpiration
Less stomata: reduces water lost through pores as there are less pores3
Leaves rolled: stomata sunken in pits that traps humid air in order to decrease the gradient for transpiration
Leaves have a waxy cuticle: reduces water loss from cell mesophyll
These help leaves exploit factors that affect plant transpiration, primarily:
1) Light intensity: It promotes photosynthesis which opens stoma
Temperature: It provides energy and favours the dark reaction
Humidity: it affects the gradient outside the leaf
Air-flow: it regulates humid air
