School Science Lessons
Plant cells and tissues
2014-12-3
Please send comments to: J.Elfick@uq.edu.au
Table of contents
9.51.0 Cells and tissues
9.52.0 Leaves
9.53.0 Roots
9.54.0 Stems
9.70.1 Plants in dry environments

9.51.0 Cells and tissues
9.9.0 Cells, plant cells, Elodea
9.0.1 Characteristics and functions of plant tissue types (Table)
9.54.2 Phloem
9.0.2 Plant tissues, plant parts
9.54.1 Xylem

Experiments
9.9.2
Plasmolysis, Elodea, Tradescantia
9.57 Cells and tissue sections, T.S., L.S., V.S., R.L.S., T.L.S.
9.56.1 Effect of temperature and chemicals on beetroot plasma membrane
9.55 Human cheek cells
9.61 Microscope staining techniques
9.9.3 Onion leaf scale cells, onion leaf epidermis, bulb
9.58 Parenchyma cells of tomato
9.59 Phloem cells of pumpkin, Cucurbita
9.54 Plant cell, cork cells, Robert Hooke
9.9.4 Plant epidermis, Tradescantia, Zebrina
9.9.5 Subsurface sections of leaf, Vinca
9.9.7 Stone cells, pear
9.60 Section cutting by hand
9.9.1 Stamen hair cells, Tradescantia
9.64 Wood cells, Eucalyptus, poplar

9.52.0 Leaves, (Experiments)
9.53.6 Cladode, Bossiaea, (GIF)
1.6 Different leaves, (Primary)
9.5.1a External features of a leaf, elm, beech, apple, Hydrangea
9.5.6 Foliage leaves, stipules
9.68 Isobilateral leaf, Eucalyptus
9.67 Grass leaf
3.27 Leaf classification, (Primary)
3.26 Leaf collection, (Primary)
9.82 Leaf of Hakea, xeromorphic leaf
5.07 Leaf, leaves
9.5.4 Leaf, stomate, apple, adaptations of stomates
9.5.5 Leaf tendrils, garden pea, sweet pea
6.6.8 Leafy crops
9.70 Leaves containing aerenchyma, water lily
9.70.1 Plants in dry environments, Acacia, Opuntia
5.31 Leaves lose water, (Primary)
9.66 Leaves of agricultural plants
9.65 Leaves of bushy plants
9.5.11 Phylloclades, butcher's broom
9.5.12 Phyllode, Acacia
9.69 Stomates in a leaf
9.66.3 Structure of dicotyledon leaf, privet, lilac
9.69.1 Structure of stomata, EucalyptusHakea

9.53.0 Roots
6.6.4 Starchy root crops
6.6.7 Tap root crops and bulb crops

Experiments
9.3.17
Adventitious roots, twig of the willow
9.73.1 Apogeotropic roots, mangroves
9.3.18 Climbing adventitious roots, English ivy
9.71.0 Dicotyledon root and monocotyledon root
9.3.7 Dicotyledon root, broad bean, buttercup
9.74 Excretion of acids by roots
9.73 Gravity affects the growth of stems and roots
9.3.6 Lateral roots, cress, coconut
9.72 Legume roots, broad bean, clover, Rhizobium
9.3.9 Mycorrhizal roots, birch, pine, heather, bird's nest orchid
5.10 Root hairs
9.3.3 Root hairs, cress
9.75 Root hairs of germinating common bean plant
6.9.16.7 Root nodules
4.3.13 Root nodules, Isolate micro-organisms from root nodules, Rhizobium
4.3.12 Root nodules, Nitrogen-fixing bacteria
9.3.15 Root pressure, Fuchsia, busy Lizzie
9.184 Root pressure, Water transport in plants, root pressure
5.10.01 Root rhizosphere
9.76 Root structure of mung bean
5.30 Roots absorb water, (Primary)
9.77 Roots absorb water, Tradescantia
1.29 Roots and stems, (Primary)
5.09 Roots, bushy plants, grasses
9.3.5 Roots, cress, mustard
9.97 Roots from plant parts
9.3.19 Specialized roots, prop roots, tap roots, tuberous roots
9.3.12 Storage roots with food reserves. potato
9.3.16 Tap roots, wallflower, groundsel
9.66.2 Young root, black mustard, white mustard

9.54.0 Stems
5.08 Woody stems, wood
9.78 Celery stalk, bean stem
9.185 Conduction of water and salts through the stems
9.6.22 Corm, false stem (pseudostem) banana, taro
9.79 Dicotyledon stem, sunflower
9.6.22 Corm, false stem (pseudostem) banana, taro
9.1.2.1 Corm, gladiolus, crocus
9.6.14 Creeping stems, moneywort (creeping jenny), ground ivy
9.6.20 Herbaceous dicotyledon stem, buttercup
9.6.6 Herbaceous dicotyledon stem, carnation
9.6.7 Herbaceous monocotyledon stem, iris
9.6.5 Herbaceous stem, forage legume alfalfa, (lucerne)
9.80 Monocotyledon stem, maize, (corn), Zea mays
9.80.1 Monocotyledon stems, Cocos nucifera coconut, Dracaena
9.1.4.1 Rhizome, ginger, iris
9.6.15 Runners, strawberry
3.28 Stems and roots, (Primary)
9.6.18 Stem hooks, bramble (blackberry), rose
9.6.16 Stolons, currant, European gooseberry, banana
9.6.13 Terminal bud, linden tree (lime tree), beech, oak
9.6.8 Xeromorphic stem, spinifex
9.6.12 Twigs of trees in winter, horse chestnut, sycamore, linden tree (lime tree), beech, oak
9.6.21 Twining stem, climbing bean, yam
9.6.17 Woody stem, hawthorn

9.54.1 Xylem
9.57 Cells and tissue sections, T.S., L.S., V.S., R.L.S., T.L.S.
9.0.1 Characteristics and functions of plant tissue types (See: Table 8. Xylem)
9.3.7 Dicotyledon root, broad bean, buttercup

9.54.2 Phloem
9.57 Cells and tissue sections, T.S., L.S., V.S., R.L.S., T.L.S.
9.0.1 Characteristics and functions of plant tissue types (See: Table 9. Phloem)
9.3.7 Dicotyledon root, broad bean, buttercup
9.59 Phloem cells of pumpkin, Cucurbita
9.63 Stem with secondary thickening, linden tree (lime tree), horse chestnut

9.0.2 Plant tissues, plant parts
See diagram 9.53: Plant parts
The tissues are formed by groups of cells that have similar or related functions. In multicellular plant bodies the cells are cemented together where adjacent cell walls touch by the middle lamella that may contain calcium pectate. Where the walls are not in contact, the spaces between them form a continuous intercellular system of air spaces. This can be very extensive in parenchyma tissues, or absent in conducting tissues. The plasmodesmata, fine cytoplasmic connections through minute pores in walls of adjacent cells, maintain continuity of the cytoplasm. When the walls become thickened, these connections are often lost. Pits, or thin areas in cell walls, are sometimes associated with plasmodesmata. In many differentiated cells, the cell walls become thick and impregnated with other substances and the protoplast disappears at maturity. These non-living units are still called cells. They function in conduction of materials (xylem) provide mechanical strength (fibres) or give protective covering (cork and bark). Tissue arrangement is different in animals where the cells have no walls and are embedded in a matrix secreted by the cells. This matrix is often extensive and lacks an air space system such as is found in plant tissues. A plant tissue may be simple or complex. A simple tissue is a group of structurally similar cells of similar origin and performing the same function, parenchyma (storage and packing) collenchyma (mechanical) sclerenchyma (mechanical). A complex tissue is a group of dissimilar cells of similar origin performing the same function, vascular tissue of the xylem containing: vessels, fibres, tracheids, and parenchyma cells.

9.3.3 Root hairs, cress
See diagram 9.75: Root hairs on germinating bean seed | See diagram 9.73.3: Root hairs in the soil
See diagram 9.73.2: Root hairs in L.S. root
1. Root hairs increase the surface area of the roots and usually occur in very large numbers. Maize,
Zea mays has 420 root hairs per mm2. Leave cress, garden cress seeds, Lepidium sativum, to swell in water in a flat glass dish for 15 minutes. Cut a square of filter paper from a sheet and wrap around a glass plate and hold in place with rubber bands. Transfer swollen cress seeds with forceps to the filter paper and put in two rows, each row 3 cm from the edge of the two narrow ends. Because their coats are mucilaginous, the seeds stick well to the paper. The glass plate is placed in a 400 mL beaker, water is added until the level almost reaches the lower row of seeds. The beaker is then covered with a Petri dish and left to stand. Within three days small cress seedlings have developed from the seeds. The roots of the bottom row that are dipping in the water have almost no root hairs. The roots of the upper row growing in air on the moist filter paper have formed a large number of hairs. The roots on the lower row have formed few roots.

9.3.5 Roots, cress, mustard
1. Soak a ceramic flower pot in water and scatter the cress seeds thinly over the inner surface. They stick to the pot because of their mucilaginous seed coats. Invert the pot over a dish containing enough water to cover the rim of the pot and put in a warm place. The seeds will quickly germinate and provide suitable roots for examination. Cut off the terminal 1 cm from the end of one root and mount in lactophenol-erythrosin. Note the arrangement of tissues at the apex, the region of elongation, the development and structure of root hairs. After making these observations, crush the specimen and examine the older region for annular and spiral vessels of the protoxylem.
2. Germinate mustard seedlings on damp absorbent paper and cover with a glass jar.
Cut transverse sections:
1. at 1 mm behind tip for apical meristematic cells and root cap cells,
2. at 2 to 4 mm behind tip for elongating cells and 3. 6 to 8 mm behind tip for elongated cells and root hairs. Stain in acid phloroglucin and mount in 50% glycerine solution. (Mustard and cress seedlings is a favourite salad sandwich filling.)

9.3.6 Lateral roots, cress, coconut
See diagram 53.5: Lateral roots of coconut
Allow cress seedlings or other seedlings to grow until the radicle shows lateral roots. Cut off the radicle just above the smallest visible laterals and mount this terminal length in lactophenol-erythrosin, coiling it round if necessary. Note the stages in the development of the lateral roots. Coconuts have no root hairs.

9.3.7 Dicotyledon root, broad bean, buttercup
See diagram 9.73.1: Root types | See diagram 9.73.2: Root L.S. | See diagram 9.73.3: Root in soil
See diagram 9.71.2: Root transverse sections, TS
1. Cut a transverse section of a broad bean root. Use the thumb and forefinger to hold the root between two pieces of pith or carrot tissue. Dip the material in water to moisten it. Never cut dry material! To cut microscope sections use a one-sided razor blade, "Gem", dipped in water. Hold the material vertically and draw the razor blade quickly across it. Cut the thinnest possible sections as wedge shapes. Wash the sections into a small dish of water. Use a camel hair paint brush to select the thinnest section, mount it in
water on a slide and cover with a coverslip. Irrigate with aniline sulfate to colour the xylem elements yellow. Look for the following tissues all embedded in parenchyma. Note the relative positions of the various tissues. Examine the tissues under high power and note the cellular structures:
1.1 the piliferous layer, with its root hairs,
1.2 the cortex, composed of the cortex proper and the endodermis,
1.3 the stele, composed of xylem (protoxylem and metaxylem) phloem and pericycle.
2. Cut a transverse section of a buttercup root. Note the creeping stems and the adventitious roots arising at nodes from stems. Observe the epidermis, wide cortex containing parenchyma, and the endodermis that is the innermost layer of the cortex. The endodermis forms a definite ring of thickened cells. See the passage cells through the endodermis opposite the protoxylem. The phloem and xylem are exarch, i.e. the first cells to differentiate are towards the outside of the stele. The xylem consists of four protoxylem groups linking with the metaxylem that occupies the centre. In the root, the first cells of the xylem to become fully differentiated, i.e. lignified, are those cells towards the outside of the core of xylem. This part of the xylem, differentiated while the root is still elongating, is the protoxylem. Metaxylem is the remainder of the xylem to differentiate after the root stops elongating. Protoxylem and metaxylem together form the primary xylem. primary xylem and primary phloem are both derived directly from the provascular tissue.

9.3.9 Mycorrhizal roots, birch, pine, heather, bird's nest orchid
Mycorrhiza roots are found growing in the surface layers of leaf mould below the trees. Note the characteristic branching. Cut sections of leaf mould to see ectotrophic mycorrhiza. Collect young, thin roots of heather in the spring. Mount a length of one of them in lactophenol-erythrosin and look for endotrophic fungal threads in the narrow cortex. Examine a section of heather root to see endotrophic mycorrhiza.

9.3.12 Storage roots with food reserves, potato
See diagram 9.85: Potato tuber
Test seeds, leaves, roots, stems, tubers for glucose and fructose.

9.3.15 Root pressure, Fuchsia, busy Lizzie
See diagram 9.184: Root pressure
Do NOT use elemental mercury for school experiments!
1. Cut a Fuchsia stem. Cut the stem of a single stem pot plant 1 cm above the soil. Fit a short piece of rubber tubing to the cut stump then fill the tubing with water. Insert into the end of the rubber tubing 50 cm of narrow bore glass tubing. Fix the glass tubing vertically and each day measure the height of liquid in it. Replace the glass tubing with a manometer and record the root pressure attained.
2. The salt concentration of the cell sap in both the root hairs and root cells is generally higher than that of their surroundings. Consequently, because of the semipermeable nature of the cell membranes, water is absorbed by osmosis through the root and rises in the plant under a certain pressure. This root pressure maintains the turgor of plant tissues. Cut a busy Lizzie or a Fuchsia, horizontally, 5 cm above the soil. Apply glycerine around the outside of the stump. Fix a piece of rubber tubing over it and bind with string. Connect a glass tube, 400 mm long, to the other end of the rubber tubing and hold it in place with a small clamp and a right angle clamp attached to a Bunsen burner stand. Attach a scale to the glass tube. Pour water into the tube so that its level can be read against the scale. Covert the tube with liquid paraffin to prevent evaporation from the surface of the water. Add water to the pot regularly and read the level of the water meniscus in the glass tube. The meniscus slowly rises in the glass tube.

9.3.16 Tap roots, wallflower, groundsel
See diagram 9.73.1: Two kinds of roots
Any grass has fibrous roots. Watercress has adventitious roots. In dicotyledons, the main axis of the root (tap root) grows vertically downwards and is continuous with the stem. It bears lateral branch roots that originate from within the root tissue. The tip of each root has a growing apex protected by a root cap. Several centimetres behind the apex is a mass of root hairs growing out from the surface layer of the root. These increase the surface area of the roots where water absorption occurs. In monocotyledons, grasses,
the fibrous root system with no obvious tap root. There are many roots of about equal size and most of them arise from the lower part of the stem at nodes (adventitious roots). The shoot system consists of a stem with long leaves attached at nodes. The leaves have no petioles, but have long sheathing leaf bases that in the young plant enfold the younger leaves. At the top of the sheathing leaf base there may be a small membranous tongue, the ligule, a common feature of grasses. The lamina is long and linear with parallel venation.

9.3.17 Adventitious roots, twig of the willow
Observe the development of adventitious roots on the twig of the willow. Gather the twigs late in the winter and place in water. In a few weeks, the buds will open and adventitious roots will appear on the stem. Keep a dated record of the progress of the shoot.

9.3.18 Climbing adventitious roots, English ivy
Detach a spray of ivy from an old wall or the trunk of a tree. Note how difficult doing this without breaking the stems of the ivy is because the adventitious roots cling to their support. Examine these adventitious climbing roots.

9.3.19 Specialized roots, prop roots, tap roots, tuberous roots
See diagram 9.87: Sweet potato tuber
1. Prop roots holds up the stem, maize (corn)
2. Swollen tap roots store food, radish
3. Tuberous roots store food, sweet potato

9.5.1a External features of a leaf, elm, beech, apple, Hydrangea
See diagram 9.66: Papaya leaf
Choose a simple form of leaf and examine its external appearance in detail. Note the leaf, showing the swollen leaf base, the petiole and the lamina. Examine the type of venation, and note how the veins gradually diminish in size, until the ultimate branches are scarcely visible.

9.5.4 Leaf, stomate, apple, adaptations of stomates
Iris and Narcissus have an isobilateral monocotyledon leaf with palisade tissue on both surfaces. The thickness of the cuticle varies on different leaves of the same plant. However, plants adapted to dry conditions, e.g. Hakea and Eucalyptus, have thick cuticles and plants growing with abundant water supply, Nymphaea, water lily, have thin cuticles.

9.5.5 Leaf tendrils, garden pea, sweet pea, (Yellow vetchling has stem tendrils.)
Note their position and relation to other leaf structures.

9.5.6 Foliage leaves, stipules
See diagram 9.66.2: Different leaves | See diagram 52.2: Breadfruit stipules | See diagram 53.3: Coconut leaf
Note the stipules of certain leaves in rose, pea. Note net venation (reticulate venation) and parallel venation in grasses. Leaves adapted for photosynthesis have the following features:
1. The broad leaf blade, lamina, has a large surface area to volume ratio and many stomata, pores, to help light absorption and gas exchange.
2. Many vein endings supply water and remove sugar from the leaf.

9.5.11 Phylloclades, butcher's broom
Note the very reduced leaves that are modified branches.

9.5.12 Phyllode, Acacia
See diagram 9.53.5: Acacia
Note the buds or branches in the axils of the phyllodes showing that these are modified leaf structures.
9.54 Plant cell, cork cells, Robert Hooke
See diagram 9.54: Plant cell
Robert Hooke, (1635-1733), examined very thin slices of cork. He noted compartments that reminded him of cells, the small rooms used by monks in monasteries. He was looking at the dead cell walls. The cavities of the compartment previously contained the living cells.
Experiment
1. Cut a wedge-shaped thin slice of cork and pith from the centre of a stem, e.g. potato, watermelon, tomato. Observe the cells as seen by Robert Hooke.
2.. Cut a wedge-shaped thin slice of a soft green stem. Look for cytoplasm, nucleus, chloroplasts, cell wall, and cell membrane.

9.9.0 Cells, plant cells, Elodea, pond weed
See diagram 9.63: Elodea cells | See diagram 9.54: Plant cell details
Plant and animal cells are similar, consisting of a protoplast bounded by a cell membrane. However, plant cells have a rigid cellulose wall surrounding the protoplast. The cell wall is in contact with its cell membrane. Mature plant cells have vacuoles and plastids. Plastids are membrane bound organelles in the cytoplasm. The three types of plastids are as follows:
1. Chloroplasts contain chlorophyll pigments and occur in all green parts of the plant.
2. Chromoplasts contain carotene and xanthophyll pigments. They give colour to all red, orange and yellow parts of the plant. The pink, purple and blue colour of plants come from anthocyanin pigments dissolved in the vacuole sap, e.g. Tradescantia, beetroot.
3. Leucoplasts are colourless plastids found in most other plant cells where starch grains may form as a storage product, potato Solanum tuberosum.
Experiment
1. Mount a complete leaf of Elodea in water on a slide and examine under high power of the microscope. Note the small green granules containing chlorophyll, chloroplasts. Observe the movement of the chloroplasts showing that the cytoplasm is moving, cytoplasmic streaming. Note the cellulose cell walls.
2. Contrast plant cells with animal cells, e.g.
9.55 Human cheek cells

9.9.1 Stamen hair cells of Tradescantia
Tradescantia, is an important plant for study because the purple hairs on the stamens are "self staining". So it is possible to observe the movements of the contents of a live plant cell with the contents already stained. Use forceps to remove a stamen from a Tradescantia young flower. Pull off one purple hair from the stamen, mount it in a drop of water, apply a coverslip and examine it under low power.
Note the following:
1. The cuticle that sheaths the cell and whole filaments of cells,
2. The cellulose cell wall around each cell,
3. The middle lamella layer between neighbouring cells only,
4. The peripheral cytoplasm containing organelles that give it a granular appearance,
5. The nucleus in the peripheral cytoplasm or suspended by cytoplasmic strands in the centre of the vacuole,
6. The vacuole containing dissolved substances, e.g. purple anthocyanin pigments,
7. The movement of granules in the cytoplasm, cytoplasmic streaming,
8. The movement of the nucleus. Check the position of the nucleus in the same cell every five minutes.

9.9.2 Plasmolysis, Elodea, Tradescantia
See diagram 9.63: Cell, waterweed, Elodea cell
See diagram 9.178: Plasmolysis in Tradescantia cells
1. Use plasmolysis to show that the cell wall is a non-living envelope distinct from the cytoplasm. Irrigate the preparation with a hypertonic salt solution more concentrated than the cell sap. By osmosis, water passes from the weaker solution to the stronger solution through the differentially permeable living cell membranes. The resulting contraction of the living cell contents is called plasmolysis. In the plasmolysed cell, the cell wall that is permeable to the solution remains rigid. The salt solution remains in the space between the cell wall and protoplast. If you then irrigate the preparation with water the protoplasts expand again quite rapidly as water passes in through the living membranes of the cytoplasm to the vacuole. The original state of turbidity of the cells returns.
2 To study the differentially permeable membranes, irrigate the preparation with iodine to kill it and again attempt plasmolysis with hypertonic salt solution. The cell no longer responds because the differentially permeable properties of the cytoplasmic membranes have died. A mature plant cell with vacuoles has two cytoplasmic membranes. The outer membrane is in contact with the cell wall. The inner membrane separates the vacuole from the cytoplasm. Use iodine solution to stain the nucleus.
3. Examine the small leaves near the end of the stem of the waterweed Elodea. Put a single small leaf in a drop of water on a glass microscope slide, cover with a coverslip and examine with a microscope. In strong light the cellular contents may have a flowing motion called cytosis or protoplasmic streaming.
4. Make a slide of living Elodea to show the presence if a cell wall. Put a drop of salt water solution on one edge of the coverslip. Draw the salt solution under the coverslip by placing a piece if absorbent towelling or blotter at the opposite side of the slip so that the liquid on the slide will rise up the paper. Water will diffuse out of the cells into the salt water. As this proceeds, the cellular contents may be observed to shrink, but the rigid cell walls retain their original structure. Other plant cells may be used to show this phenomenon. Fleshy leaves with a thin layer that can be peeled off are possible sources of thin cellular layers.
5. Repeat the experiment with Tradescantia, lettuce and spinach cells.

9.9.3 Onion leaf scale cells, onion leaf epidermis, bulb
See diagram 9.56: Plant cell (diagrammatic) | See diagram 2.30: Detach epidermis from leaf
A. cell wall, B. middle lamella, C. nucleus, D. cytoplasm, E. plasma membrane, F. tonoplast (plasma membrane) G. vacuole containing cell sap
1. An onion bulb is a condensed shoot with a very short stem enclosed by fleshy leaf bases, leaf scales. Cut an onion bulb in half, longitudinally (downwards). Use forceps to peel off the thin epidermis from the concave (inner) side of an onion leaf scale. Put a small flat piece of it in a water drop on a microscope slide. Apply a coverslip and examine the structure of the cells under low power. Stain the cell contents by putting one drop of iodine solution at the edge of the coverslip, then draw the solution under the coverslip by putting absorbent paper on the opposite edge. Note the cytoplasm enclosing several large vacuoles and the normally colourless nucleus now pale yellow because of the iodine solution. Note the thin cellulose cell wall surrounding the whole cell.
2. Methyl green acetic acid solution and carmine acetic acid solution simultaneously fix and stain. Transfer a drop of methyl green acetic acid to a slide with a glass rod. Detach a small piece of epidermis from the inner side of a scale of an onion. Put it immediately into the drop of methyl green acetic acid. Apply a coverslip and examine the preparation at a magnification of 250 X. The cell nuclei will be stained a strong blue-green colour, while the cell walls will only be weakly tinted. The rest of the cell contents remain
unstained. The image in the microscope will be even more contrasting if, after the desired intensity of staining has been reached, the dye solution is replaced by 2% acetic acid. Apply a drop of 2% acetic acid to one edge of the cover glass with a glass rod, and suck it under the cover glass by applying a piece of filter paper to the opposite side. If carmine acetic acid is used, the cell nuclei are stained a deep red. Use methyl green acetic for more fragile plant specimens and for showing the nuclei of protozoa.

9.9.4 Plant epidermis, Tradescantia, Zebrina
See diagram 9.69.3: Stomate, surface view, guard cells | See diagram 9.65.3.1: Section view of a leaf
See diagram 9.69: Stomate guard cells
1. Use a razor blade to make an incision on the under surface of a Tradescantia leaf and strip off a small section of epidermis. Mount outer surface uppermost in water. Tradescantia has no stomata in the upper epidermis.
Observe the following:
1.1. The guard cells that should be open if the weather is bright.
1.2. The extra thickening on the walls of the guard cells is next to the pores.
1.3. Accessory cells surround the guard cells.
1.4. The larger epidermal cells have colourless plastids. Guard cells have chloroplasts but epidermal cells do not. Leucoplasts (colourless plastids) cluster around the nucleus of the accessory cells.
2. Plants possess a skin called the epidermis. To study the epidermis of the leaf of a trailing plant, transfer a drop of water from a beaker to a microscope slide with a glass rod. Stretch a trailing plant leaf, with the lower surface facing upwards, over the index finger of the left hand, holding it in place with the middle finger. Make a small cut on the surface of the leaf with a dissecting needle. Grasp the torn edge with pointed forceps and peel off a small piece of the epidermis of the lower surface of the leaf. Put it, with the external surface upwards, in the drop of water on the slide. Mount a coverslip. Examine the slide under low power. Note the shape of the epidermal cells and the bean shape cells that always lie together in pairs, called "guard cells". A stomate is this pair of cells and the opening between them. Most of the water lost by the plant, and all the exchange of gases takes place through these stomates. The guard cells of the stomates may be surrounded by four cells that differ from that of the other epidermal cells. This group of six cells is called a stomatal apparatus. Find a stomatal apparatus. Examine the slide under high power. Note the stomatal apparatus and the epidermal cells surrounding it. Note which epidermal cells contain chlorophyll grains (i.e. have chloroplasts). Observe the cell nuclei.

9.9.5 Subsurface sections of leaf, Vinca
Use a variegated leaf of Vinca to cut thin subsurface sections just below the upper and lower epidermis. To do this, hold a leaf over your index finger and anchor it with your thumb and third finger.
Observe the following:
1. The upper epidermal cells and the tops of the palisade cells. The air spaces between palisade cells are small in diameter but extend vertically through the tissue. Mount this section with epidermis uppermost.
2. The lower epidermal cells and part of spongy mesophyll cells with large air spaces. Mount this section with the spongy mesophyll uppermost.

9.9.7 Stone cells, pear
See diagram 9.78.4: Stone cells
Pull apart the gritty tissue of pear fruit on a slide. Examine the structure of the stone cells. Note the absence of cytoplasm and look for simple unbranched and branched pits. Stain with iodine solution or aniline sulfate or aniline chloride to show the lignified walls.

9.55 Human cheek cells
See diagram 9.3.67: Human cheek cells. Note the nucleus, cytoplasm, plasma membrane or plasmalemma, and granules.
You may have to seek approval to do this experiment because saliva can carry disease. Instead of taking cheek cells you can use prepared slides of cheek cells from a school laboratory supplier.
Cheek cells come from the stratified squamous epithelium tissue on the surface of the mucous membrane inside the cheek. These flat, scale-like cells are shed constantly as the tissue is renewed so it is easy to obtain some for study by gently scraping the inside of the cheek. This tissue is not keratinized so the surface cells are still living and have live nuclei, in contrast with shed epidermal cells. Similar tissue lines the vagina.
Experiment
1. Observe human epithelial cells from inside the cheek. Use a clean toothpick to gently scrape the inside surface of the cheek. Put the whitish scraping into a drop of water or 0.65% saline solution on a microscope slide. Add a drop of stain, e.g. methylene blue or iodine solution, and apply a coverslip. View under low power and high power. Note the protoplasm containing a central nucleus and granular cytoplasm. The outer boundary of the protoplasm is the plasma membrane. Adjacent cells look like paving stones. The
nucleus and cytoplasm have a different refractive index, so note the interfaces between them. In later experiments, compare the animal cell with the plant cell. The animal cell has no cell wall.
2. Contrast animal cells with plant cells, e.g.
9.9.0 Cells, plant cells, Elodea

9.56.1 Effect of temperature and chemicals on beetroot plasma membrane
This experiment is an indirect study of the effects of different substances and treatments on living beetroot cells. Each beetroot cell has a large central vacuole bounded by a plasma membrane. The vacuole contains the red pigment anthocyanin that gives the beetroot its typical colour. The beetroot cell is also surrounded by the cell membrane. If both membranes remain intact, the anthocyanin cannot escape into the surrounding environment. If both membranes are stressed or damaged, lysis occurs and the red colour can leak out. The cell wall surrounding plant cells provides a structure to the plant. It does not have a role in controlling the movement of substances into and out of cells. The dependent variable is the colour of the beetroot cells. The manipulated variables could be temperature, or the concentration of chemicals, e. g. alcohol.
If you have a spectrophotometer the ?max = 535 nm.
What is the maximum temperature a cell can sustain before lysis occurs? How does alteration of temperature impact on the structures of a cells? How can the addition of alcohol or saline chemicals to the environment impact on the cell membrane stability?

9.57 Cells and tissue sections, T.S., L.S., V.S., R.L.S., T.L.S.
See diagram 9.57: Tissue sections 1 | See diagram 9.57.1: Tissue sections 2
See diagram 9.57.2: Wood section
A slice across a stem, at right angles to the axis of the stem, is a transverse section, T.S. Any slice parallel to the axis of a stem is a longitudinal section, L.S. A slice vertically down is a vertical section, V.S. A slice parallel to the axis of the stem, along the radius, is a radial longitudinal section, R.L.S. A slice parallel to the axis of the stem, along a tangent to the cross-section, is a tangential longitudinal section, T.L.S.
Experiment
1. Cut a wedge-shaped transverse section across a soft stem, e.g. tomato, potato, sunflower. Note the groups of similar cells, tissues. The epidermis is the one cell thick outer layer. It may have a waxy cuticle on the outside to protect against desiccation. The bundles of cells, vascular bundles, contain food conducting phloem cells on the outside and water conducting xylem cells on the inside. The walls of the xylem cells, vessels, are strengthened. Old xylem forms wood. Groups of cells with very thick walls, sclerenchyma, strengthen the stem. Parenchyma tissue is the loose packing cells. Between the xylem and the phloem are closely packed cells with large nuclei and thin walls, the cambium. Cambium cells produce new cells by mitosis to make the stem thicker. Draw a map diagram to show the different tissues.
2. Observe the remaining stump of a cut down tree or the sawn end of a thick branch. Note the sap wood, heart wood, annual rings, phloem and bark. The appearance of the rays shows the type of section. In transverse section, T.S., the rays are radial lines often only one cell in width. In radial longitudinal section, R.L.S., the rays appear as partial brick walls. Any broken appearance is caused
by the section not being exactly radial. In tangential longitudinal section, T.L.S., the rays appear as lens-shaped areas and from this type of section the actual vertical extent and width of the rays may be accurately determined. L.S., longitudinal section, refers to any section at right angles to the axis. Examine the T.S., R.L.S. and T.L.S. sections of the wood of the linden tree (Tilea europea). Find
the rays and identify the type of section.

9.58 Parenchyma cells of tomato
See diagram 9.58: Parenchyma cells of tomato
1. Look for the thin cell wall, plasma membrane, vacuole, cytoplasm, chromoplasts, and nucleus.
2. Remove a very small portion of the pulpy tissue immediately beneath the skin of a tomato fruit. Mount this on a slide in water and then tease it out with dissecting needles. Apply a coverslip and examine under high power. Note the parenchyma cells containing orange-red chromoplasts and cytoplasm, nucleus and vacuoles. Stain with iodine solution and examine the structure in detail.

9.59 Phloem cells of pumpkin, Cucurbita
See diagram 9.59.1: T.S. Pumpkin stem | See diagram 9.59.2: T.S. Vascular bundle
See diagram 9.59.3: L.S. and T.S. phloem cells, high power
1. Note the collenchyma, cortex, endodermis, pericycle, pith (often broken), parenchyma (packing tissues), phloem, xylem, cambium, bicollateral vascular bundle (phloem both outside and inside xylem), characteristic of pumpkins and melons.
2. Examine a transverse section and a longitudinal section. Note the general arrangement of the bicollateral bundles, with the phloem both internal and external to the xylem in the vascular bundles. Sieve tubes form vertical files of cells placed end to end. Where each cross wall is perforated is called a sieve plate. Each sieve tube element has a companion cell next to it. Companion cells are small with dense cytoplasm.

9.60 Section cutting by hand
See diagram 9.60: Cut sections by hand
1. Use a razor blade, preferably one-sided, to cut a very thin slice from a cork or a stick of pith.
Be careful! Cut away from the body! Examine the slice with a magnifying glass or low power microscope. Cut an incomplete shaving, like a thin slice of cheese, and examine its thinnest edge. Note the arrangement of the dead cell walls.
2. Cut a transverse section, T. S., at right angles to the long axis of the organ or plant. Cut a longitudinal section, L. S., parallel to the axis of the organ or plant. Cut a radial longitudinal section, R. L. S., as with a longitudinal section but cut along the radius of the organ or plant.
3. Make a transverse section by cutting a carrot or piece of pith in half longitudinally. Then hold the tissue to be sectioned between the two halves of the carrot or pith and cut across, away from you, with a one-sided razor blade, e.g. "Gem".

9.61 Microscope staining techniques
See diagram 2.26: Drawing stain across specimen under coverslip
3.13.1 Haematoxylin solution, Delafield's haematoxylin, microscopy stain
Use safety glasses and nitrile chemical-resistant gloves when working with stains.
1. Irrigation
Mount a section of plant tissue in a drop of water on a microscope slide. Put a coverslip on the drop of water so that no air bubbles remain under the coverslip. Put a drop of stain near the edge of the coverslip so that it is in contact with the edge of the drop of water. Touch the other side of the drop of water under the coverslip with absorbent paper to draw the stain across the plant tissue.
2. Immersion in iodine solution
Put sections of plant tissue in iodine solution for one minute. Remove the sections, rinse in tap water and mount them on a microscope slide in dilute glycerine.
3. Immersion in acid phloroglucin solution
Put sections of plant stem in phloroglucin solution for 30 seconds. Use a mounted needle to transfer the section to a drop of concentrated hydrochloric acid for two seconds. Mount the section in glycerine and apply a coverslip. Observe the lignin in xylem or woody tissue stained bright red.
4. Immersion in safranin and haematoxylin solutions
Put sections of plant tissue in 50% by volume alcohol / water solution. Use a mounted needle to transfer sections to safranin solution. Wash sections with tap water. Transfer sections to 's haematoxylin solution. Observe the section under a microscope to monitor the staining. If the section is overstained, destain in acidified alcohol solution. Wash sections, mount in glycerine and apply a coverslip.

9.63 Stem with secondary thickening, linden tree (lime tree), horse chestnut
See diagram 9.57.4: Tilia TS Stem
1. Examine transverse sections through the stem of various ages as follows:
1.1 near the apex of the shoot,
1.2. the middle of the first year's growth,
1.3. near the bottom of the first year's growth,
1.4. about the middle of a later year's growth.
Note the secondary wood and examine the vessels of the spring and autumn wood. Look for medullary rays. On the outside of the cambium, note the secondary phloem, containing thickened cells called phloem fibres. Note how the medullary rays widen out in the phloem. Near the periphery, look for cork cambium, and note the layers of cork cells produced on the outside of this.
2. Examine a secondarily thickened stem by means of radial longitudinal and tangential longitudinal sections.
Observe the following tissues:
2.1 The periderm consists of phellem and phellogen. Phellem (cork) cells have radial rows of cells with suberized walls, formed by division of the phellogen (cork cambium) one row of radially flattened cells with thin walls. In some plants the phellogen may also produce a farther layer, the phelloderm, towards the inside.
This layer is not apparent in Tilia. The lenticels, part of periderm, are regions of rounded, loosely packed cells, which allow exchange of gases through the otherwise impermeable tissue
2.2 The secondary phloem, in wedges, consists of alternating bands of fibres and sieve tubes, companion cells and parenchyma.
2.3 The cambial zone
2.4 The secondary xylem, wood. (Greek: xylon, wood)
2.5 Primary rays extend from the cortex to the pith and are very wide in secondary phloem.
2.6 Secondary rays are in secondary xylem and phloem and do not extend to the centre or to the cortex.
2.7 Primary xylem surrounds the medulla. The interpolation of secondary vascular tissues between primary xylem and primary phloem creates considerable stress in the stem.
3. The medulla and primary xylem are least affected. In the outer layers, accommodation to the increasing circumference is accomplished by the following changes:
3.1 The epidermis initially keeps pace with growth by radial cell divisions but eventually is replaced by a periderm of superficial origin, usually just beneath epidermis.
3.2 The cortex increases in circumference by expansion of cells in the tangential plane, and divisions in the radial plane. Parenchyma is mostly squashed, but collenchyma retains its form and the new walls from recent cell divisions are evident.
3.3 Parenchyma cells of the primary rays between phloem wedges expand and divide similarly. Some of these rays become conspicuous by great increase in width towards the periphery.

9.64 Wood cells, Eucalyptus, poplar
See diagram 9.57.2: Section of cut wood
Be careful! Do not allow students to use concentrated nitric acid. Use safety glasses and nitrile chemical-resistant gloves when working with concentrated acids.
Prepare woody elements for microscopic examination by maceration of a small woody twig. In a fume cupboard, fume hood, cover the pieces with concentrated nitric acid, add crystals of potassium nitrate, and heat the beaker in a fume cupboard. When the reaction has finished, remove all the acid by repeated rinsing with water. Mount twig tissue in 50% alcohol / water mixture and tease it apart with mounted needles. Observe vessels with characteristic thickening on the walls, wide lumens (internal spaces) and perforated end walls. Observe xylem parenchyma fibres and tracheids, long narrow cells with lignified walls and narrow lumen.

9.65 Leaves of bushy plants
See diagram 9.65.1: Parts of a leaf | See diagram 9.65.2: Mung bean leaves
9.65.3.1: VS of leaf showing water movement
1. Examine the leaves of a bushy plant. Most leaves are flat and thin to catch plenty of sunlight.
The bushy plant leaf has the following 3 parts:
1.1 lamina, 1.2 petiole, 1.3 leaf base.
The leaf base attaches the leaf to the stem. The petiole turns and holds up the leaf blade, like a handle. The leaf blade takes in sunlight to make food. Leaf veins are arranged in a network. A leaf is attached to the stem. In the angle between the leaf and stem is the axillary bud. A leaflet is part of a leaf. There is no axillary bud between the leaflet and the stem.
Choose a simple form of leaf and examine its external appearance in detail. Note the leaf, showing the swollen leaf-base, the petiole and the lamina. Examine the type of venation, and note how the veins gradually diminish in size, until the ultimate branches are scarcely visible.
Cut a vertical section of a leaf or examine the tissues in a prepared slide.

9.66 Leaves of agricultural plants
See diagram 9.66: Papaya leaf
Examine leaves of agricultural plants, e.g. banana, breadfruit, chilli, cassava, cocoa, coconut, pineapple, swamp taro, sweet potato, yam. Draw each leaf and label the parts and describe the leaf in your own words. For example, the lamina of the leaf of the Papaya plant has tooth-shaped lobes and the petiole is long and hollow.

9.66.2 Young root, black mustard, white mustard
Cultivate mustard seed on damp absorbent paper. Use a hand lens or low power of a microscope to see the cylindrical radicle, the root cap and the root hairs. Cut off the radicle then cut it longitudinally down the middle and mount in water. Observe a single mature hair. It is an outgrowth of an epidermal cell.

9.66.3 Structure of dicotyledon leaf, privet, lilac
See diagram 9.65.7: Privet leaf
1. Examine leaves on a light table. Draw the outline of the leaf and include the mid rib and veins.
2. Choose a part of a leaf that contains some of the midrib. Fix it between two pieces of elder pith or carrot so that you can cut the midrib transversely. Mount the section and stain with an aniline salt. Examine the structure under the low power, noting the upper epidermis covered with a layer of cuticle, palisade tissue, spongy tissue, and lower epidermis covered with a slightly thinner layer of cuticle. Note also the vascular bundle forming the midrib, composed chiefly of xylem and phloem. Note the midrib
embedded in a sheath of parenchyma cells and the thin portion of the leaf. Note the absence of chloroplasts in the upper and lower epidermis, a large number in the palisade mesophyll cells and a relatively smaller number in the spongy mesophyll cells. Note the shape and position of the chloroplasts. Examine the shape of the cells of the palisade tissue and note the number of layers here. The cells are separated by small air spaces. Note the irregular shape of the spongy mesophyll cells and the air spaces between them. Privet is a mesophyte dicotyledon with dorsi-ventral leaves.
3. Observe the following:
1. The upper epidermis with stomata, cuticle and glandular hairs.
2. The palisade mesophyll consisting of vertically elongated cells with chloroplasts.
3. The spongy mesophyll consists of loosely packed cells with air spaces and chloroplasts.
4. The lower epidermis has stomata, cuticle and glandular hairs.
5. Small lateral veins, often cut obliquely during preparation of the microscope slide, are situated between the palisade and spongy mesophyll.
6. The midrib is a continuation of vascular tissue of the leaf stalk. In the midrib vein, the xylem is uppermost and the phloem is on the underside.

9.67 Grass leaf
See diagram 9.67: Grass leaf
The grass leaf has three parts: 1. leaf blade 2. leaf sheath and 3. ligule, an outgrowth where the leaf blade and sheath join. The leaf veins are arranged in parallel.
Cut a vertical section of a grass leaf or study a prepared slide and observe the following:
1. upper and lower epidermis, chlorenchyma,
2. lack of differentiation into palisade and spongy mesophyll,
3. small lateral veins cut at right angles because of parallel venation,
4. sclerenchyma forming L-shaped girders around lateral veins and midrib,
5. border parenchyma surrounding the veins.

9.68 Isobilateral leaf, Eucalyptus
See diagram 9.65.8: Eucalyptus, T.S.
When Eucalyptus leaves are isobilateral, twisting of the petiole allows the leaf to hang vertically and show many xeromorphic characters, e.g. thick cuticle and sunken stomata. Observe the mesophyll with palisade cells, like fence palings, next to both surfaces and the oil glands seen as large circular cavities.

9.69 Stomates in a leaf
See diagram 9.69: Stomate, T.S. and V.S. | See diagram 9.69.1: Surface view of the lower epidermis of Kalanchoe
1. Use a one-sided razor blade to make an incision on the lower surface of a soft leaf and use forceps to strip off a small section of epidermis. Be careful! Cut away from the body! Mount the strip in water on a microscope slide with the outer surface uppermost. Most soft leaves have no stomates in the upper epidermis. A stomate is a small pore surrounded by two kidney-shaped guard cells. The guard cells usually contain s but epidermal cells do not usually contain chloroplasts. Put salt solution on the stomate and see if the guard cells become plasmolysed and so close the pore of the stomate. Plasmolysis is the contraction of the protoplasm away from the cell wall due to loss of water through osmosis. The stomate should be open if the weather is bright and sunny.

2. Cut a transverse section of the leaf and look for a stomate cut in section. Notice the shape of the guard cells and the pore. Note also the large air space in the mesophyll immediately adjoining the stomate pore.

3. Pour collodion on the lower surface of a leaf. Wave the leaf in the air until the collodion dries, then pull it off as a strip. See and feel the shape of the stomates in the leaf.

4. Note the distribution of stomates on leaves of Tradescantia. Use a glass rod to transfer one drop of water from a beaker on a microscope slide. Detach a leaf from a hanging plant, stretch it over the index finger of the left hand, outer surface upwards, holding it in position with the thumb and middle finger. Make a scratch along the surface of the leaf with a dissecting needle. Grip one edge of the scratch with a pair of pointed forceps and detach a small section of the outer skin, epidermis, from the upper surface of the leaf. Place it with the outside surface facing upwards in the drop of water on the slide and put a cover slip over it. Using the same technique, prepare a specimen of the outer skin, epidermis, of the underneath surface of the leaf of the same plant. Examine both preparations under low power. Note whether stomates are distributed in similar numbers on the upper and lower surface of the leaf. Examine three different areas of each specimen. Count how many stomates can be seen in the microscopic field of vision in each case and calculate the mean values. Compile a table of results.

5. Note the distribution of stomates on leaves of iris. Use the same technique to prepare a specimen of the outer skin, epidermis, of both sides of an iris leaf. Examine the specimens under low power. Note the distribution of stomates on the different sides of the leaf. Examine three different areas of each specimen as above. Calculate the mean values.

6. Note how the stomates are distributed on either side of the following: a Tradescantia leaf, an iris leaf. Note whether the iris leaf has an upper and a lower surface. Examine the stomates of the iris leaf under greater magnification. Compare their structure with that of the stomates of the hanging plant, and note whether both structures are identical. Investigate the distribution of stomates on both sides of the leaves of other terrestrial plants, e.g. Eucalyptus, an isobilateral leaf.

7. Investigate the distribution of stomates and its relation to the plant environment. The stomates are responsible for most of the water diffusion and gas exchange that occur in a plant. So the distribution of stomates on the leaves is adapted to the environment of the plant. Investigate the distribution of the stomates on both sides of a water lily leaf and a leaf of curled pond weed.

8. Put a drop of water on a microscope slide. Stretch a water lily leaf with its outer surface upwards over the index finger of the left hand, holding it in position with the thumb and middle finger of the right hand. Make a scratch along the surface of the leaf with a dissecting needle. Be careful! Do not press too hard and puncture through the leaf into the skin. Grip one edge of the scratch with a pair of pointed forceps and detach a small piece of the epidermis from the upper surface of the leaf. Put the piece of epidermis in the drop of water on the slide, with the outside surface facing upwards and put a coverslip over it. Use the same technique to prepare a specimen of the epidermis, of the underneath surface of a water lily leaf. Examine both preparations under a microscope. Compare how the stomates are distributed in the upper and lower sides of the water lily leaf.
9. Use the above technique to prepare a piece of the epidermis from both sides of a leaf of curled pond weed. Examine these preparations under low power.
10. Stomates control the movement of gases in and out of a leaf, making carbon dioxide available for photosynthesis, and controlling the loss of water from the leaf through transpiration. Stomatal density varies between monocotyledons and dicotyledons, between plant species, and between the underside and top side of the leaves on a plant.
Measure stomatal density
1. Use clear nail varnish to make an impression of the epidermis, Germolene New Skin, water-based varnish
The image shows a surface view of the lower epidermis of Kalanchoe (Kalanchoe sp.), a dicotyledonous plant. Three stomata and their associated guard cells are shown.
Each pore or stoma is surrounded by two sausage-shaped guard cells, which change shape to control the size of the stomatal aperture. In the majority of leaves with an upper and lower surface (dorsiventral) like this dicot, most stomata occur in the lower epidermis. They are usually evenly distributed in the leaves of monocots.
The stomata of most species open in daylight and close in the dark. Those plants that use CAM photosynthesis (an adapatation to reduce water loss in arid conditions), stomata close during the heat of the day, to reduce evapotranspiration, and open at night to absorb carbon dioxide for use in photosynthesis.
The guard cells contain chloroplasts (visible in this image), but in most plant species they are not able to carry out the full process of photosynthesis.
The wavy blue lines, looking rather like a jigsaw puzzle, are the cell walls of the epidermal cells. Guard cells develop and differentiate from epidermal cells.
Selecting your plants
One of the best plants for doing epidermal peels is the red hot poker plant Kniphofia. Being a monocot its stomata are highly ordered in rows, but they are big and great for stomatal opening and closing using solutions of different concentrations.
Almost as good is the Elephants Ear Saxifrage Bergenia. This also peels very easily, but the stomata are smaller although clearly visible at x100 magnification. This is a dicot so the distribution is more random.
Geraniums (a dicot) and spider plants (another monocot) also make excellent stomatal peels.
Using Clear Nail Varnish
Using clear nail varnish is a traditional method to measure stomatal density, since making the impression and viewing it under a microscope can be completed in one lesson. However, some leaves are prone to damage from the solvent in the nail varnish. The leaves absorb it, turn brown, and fail to produce any impression. Pupils lose interest and get frustrated because their leaves 'aren’t working'. Also, for a GCSE class, several pots of nail varnish are needed so that no one is left waiting, thus adding to expense.
•Prepare an epidermal impression by coating the leaf surface with nail varnish. Peel off the dried layer of nail varnish by using sellotape and stick this onto a slide.
•Alternatively, with some plants you can peel off an epidermal strip directly, which you can mount in water on a slide and place under the microscope.
•If you have an eyepiece graticule which you can use, you can work at a relatively low power, and you can count the number of stomata within different squares to act as replicates.
• If you do not have an eyepiece graticule, you can work at a higher magnification and count a number of different fields - the area visible under the microscope - at any one time.
Make sure that:
1. You get enough counts to be able to analyse your results statistically,
2. You calculate the area of leaf which you are counting in order to give a quantifiable result e.g. stomata per square mm. You will need to calibrate the size of the field of view, or the size of individual squares within a field, using a stage micrometer to do this.
Using 'New Skin'
A good alternative is to use Germolene 'New Skin' to take the impressions. There is an excellent protocol on Measuring Stomatal Density on the Practical Biology website.
Using Water Based Varnish
A successful alternative to nail varnish is to use a clear water based varnish (available at 'Do it Yourself' supermarkets). A half litre tin is cheap, and can be divided up into smaller amounts for ease of use. Paint the opaque varnish thinly on to the leaf to produce a clear film. Leave it to dry as usual. These water based varnishes take longer to dry, so if the leaves are coated during one lesson, the impressions can be peeled off and examined the next. The varnish is non toxic, so can be used on living plants without removing the leaves – this means that school plants do not have to be denuded for this experiment! In addition to revealing the stomata, the cell walls also show up.
Other Methods
Other suggestions include producing impressions on acetate film, by placing a leaf in propanone and then pressing it onto the acetate. This does not work for some plant leaves, especially those that have an uneven surface – and the leaf still has to be removed from a plant. Another method is to rub a board pen over the surface. The solvent-based ink permeates the leaf, showing up the stomata. However, this seems to work only with certain types of pen – probably related to the strong solvent in the pen. This also raises health and safety issues.
Suggestions for investigations
As well as studying stomatal patterns and densities in a variety of plants, the following questions may be posed to students:
•Does the density vary over a leaf surface?
•Does the density vary between different leaves of the same plant? . . . or between different plants of the same species? (I have looked at a number of the Brassicaceae: B. oleracea v. capitata (cabbage), v. gemmifera (brussels sprout), v. italica (broccoli), v. botrytis (cauliflower). All these leaves are available from greengrocers - and you can eat the rest of the vegetable afterwards!
Does the density vary between plants from different habitats. (I have used a number of cacti and succulent plants to do this.)
•For plants that reproduce vegetatively, is there any difference between parent and offspring? (Kalanchoe and begonias are useful here).
With a reasonable microscope it should be possible to count the stomata whilst still intact on the leaf if you have your lighting right. Depending on the species the cells can be quite obvious. Then it’s a matter of picking a small area or areas and counting. A graph grid printed on a transparency works really well. This will give you perhaps a per centimetre squared ratio and if you like extrapolating the data to get a per leaf ratio and then a comparison based on total surface area for a particular species dependant on foliage density.
Hope it helps, its good science.
We’ve connected a microscope to a TV screen and had the student calculate the actual area (taking into account the magnification etc) and then being on a TV screen she was able to easily count each stoma (I think the student even ‘sticky noted’ each stomata so she counted correctly).  We used this as an approximation of stomata per unit squared.

9.69.1 Structure of stomata, Eucalyptus, Hakea, iris, privet, narcissus, water lily
See diagram 9.65.3.1: VS Leaf | See diagram 9.69: Surface view and section view of a stomate
See diagram 9.69.3 Surface view Guard cells | See diagram 9.69.1: Hakea stomate
1. Chose a small portion of the leaf, and tear off the epidermis as a thin layer. Mount the piece of epidermis flat in water and examine it under high power. Examine a stomate and surrounding cells as a small pore surrounded by two kidney shape guard cells. The guard cells usually contain chloroplasts but epidermal cells do not usually contain chloroplasts. While looking at a stomate irrigate with salt
solution and see the guard cells become plasmolysed closing the pore of the stomate.
2. Cut a transverse section of the leaf and look for a stoma cut in section. Notice the shape of the guard cells and of the pore itself. Note also the large air space in the mesophyll immediately adjoining the stomate pore.

9.70 Leaves containing aerenchyma, water lily
See diagram 9.70: T.S. Water lily leaf
Leaves of water lilies float on the surface of the water so xylem and mechanical tissues are reduced and aerenchyma is typically present. Stain a section in acid phloroglucin solution and mount it in glycerine.
Observe the following parts:
1. Upper epidermis with cuticle and stomata,
2. lower epidermis without cuticle,
3. palisade mesophyll interspersed with elongated sclereids,
4. aerenchyma containing large intercellular cavities and interspersed with stellate sclereids,
5. veins showing small amounts of xylem.

9.70.1 Plants in dry environments, Acacia, Opuntia
(9.70.1: Acacia, wattle, Opuntia, prickly pear) | (9.5.12 Phyllode, Acacia) | (9.69.1 Structure of stomata, Hakea) | 
(9.6.8 Xeromorphic stem, spinifex)  |(9.82 Leaf of Hakea, xeromorphic leaf ) | (9.73.1 Apogeotropic roots of mangroves)
Plants growing in places of low rainfall, shallow sandy soil that will not hold water for long have adaptations in their leaves to cut down the loss of water or to store water.

9.71.0 Dicotyledon root and monocotyledon root
See diagram 9.71.1: T.S. Young root with root hairs
See diagram 9.71.2: T.S. Older root, high power (not the same plant)
The xylem vessels carry water and dissolved substances from the root up towards the shoot. The sieve tubes in the phloem carry water and dissolved foods towards the root. The xylem is arranged in a star-shaped pattern with 5 or 4 points. Small protoxylem vessels are at the points of the star outside the larger metaxylem vessels. Between the points of the xylem star are groups of sieve tubes and companion cells of the phloem. The xylem and phloem are surrounded by parenchyma tissue. Within the parenchyma are meristematic cells of the root cambium that later produce secondary thickening of the root. The xylem, phloem and parenchyma and the layers of parenchyma cells surrounding the pericycle, together are called the stele or vascular cylinder. The outer layer of the vascular cylinder is the pericycle that later forms fibres. The cortex is the outer cylinder of parenchyma and intercellular spaces. The innermost layer of the cortex forms the endodermis, a layer of cells that controls movement of solutions into and out of the stele. Later, the endodermis has suberized thickening but cells opposite the protoxylem groups, remain thin-walled and are called passage cells. The outermost layer of the root is the piliferous layer that produces root hairs. Lateral roots originate in the pericycle. Monocotyledon roots have much pith and many scattered xylem and phloem bundles.

Experiment
Cut a transverse section of a broad bean root. Use the thumb and forefinger to hold the root between two pieces of pith or carrot tissue. Dip the material in water to moisten it. Never cut dry material. To cut microscope sections use a one-sided razor blade dipped in water. Hold the material vertically and draw the razor blade quickly across it away from the body. Cut the thinnest possible sections as wedge shapes. Wash the sections into a small dish of water. Use a camel hair paint brush to select the thinnest section, mount it in water on a microscope slide and apply a coverslip. Irrigate the section with aniline sulfate to colour the xylem elements yellow.
Observe the following:
1. The piliferous layer, with its root hairs,
2. The cortex, composed of the cortex proper and the endodermis,
3. The stele, composed of xylem (protoxylem and metaxylem) phloem and pericycle. These tissues are all embedded in parenchyma.
4. Note the relative positions of the various tissues. Examine the tissues under high power and note the cellular structures.

9.72 Legume roots, broad bean, clover, Rhizobium
See diagram 9.72: Legume root and root nodules | See diagram 9.209: T.S. Root nodule
See diagram 9.72.1: Winged bean, pigeon pea, mung bean
Look for root nodules on legumes, clover. Prepare a transverse section of such a root to pass through a nodule. Note the red colour usually shown by the central part of the nodule and study the large infected cells present in this region.

9.73 Gravity affects the growth of stems and roots
1. Sprout seeds and select one that is straight. Pierce the seed with a long pin or needle and stick this into a cork. Put damp cotton or absorbent paper in a bottle. Put the cork and seedling in the bottle. Put the bottle in dark cupboard and look at it every four hours.
2. Put seeds that grow rapidly, oats, radish, or mustard seeds on moist absorbent paper between two glass plates secured with rubber bands. After germination, turn the apparatus vertically through 90o and leave to remain undisturbed. Repeat the turning at intervals and observe the effect on the roots.
3. A technique used in some laboratories is to put developing plants in a shaking machine. Without a sense of gravity the plants the tissue, e.g. orchid tissue plantlets, does not differentiate into roots and shoots. This process allows undifferentiated tissue to grow bulkier and prepare for transplanting.

9.73.1 Apogeotropic roots of mangroves
See diagram 9.73: Mangrove roots
Observe mangroves in tidal swamps. They have apogeotropic roots containing aerenchyma. The roots grow upwards. Aerenchyma gas spaces provide an internal passage for oxygen gas in plants growing in flooded and anaerobic habitats.

9.74 Excretion of acids by roots
See diagram 9.74: Excretion of acid by roots
Plants excrete acids through the roots. These acids dissolve the otherwise insoluble chalky constituents of the soil. Put a marble plate or bathroom tile with the polished side upwards in a sloping position in a flowerpot. Fill the flowerpot with soil. Plant a bean seedling with roots about 2 cm long in such a position that the roots are forced to grow along the polished surface. After three weeks, remove the marble plate and rinse it with water. The polished surface of the marble plate has become etched where it was in contact with the roots. To make the etched lines clear, apply black shoe polish with a pad of cotton wool. The acids excreted by the roots of the bean plant have dissolved the marble, calcium carbonate, at the points of contact.

9.75 Root hairs of germinating bean plant
See diagram 9.75: Root hairs of germinating bean seed
Put bean seeds or mustard seed on damp absorbent paper. Cover the seeds to keep the paper damp. After germination, use a magnifying glass to examine the cylindrical radicle, the root cap and the tiny root hairs growing from the side of the root just behind the root tip. They are very thin walled outgrowths of the epidermal cells. Most plants take water and plant nutrients into their roots through the root hairs. Root hairs may be damaged by careless transplanting, salty soil and lack of oxygen in waterlogged soil.

9.76 Root structure of mung bean
See diagram 9.72.1: Mung bean plant
Wash the soil from the roots of a small bushy plant, e.g. mung bean, and a grass, e.g. para grass. Bushy plants have a main root, the tap root or primary root, and smaller lateral roots or secondary roots. These roots can grow very deep. Grasses and palms have no main root, only many fibrous roots. These are thin roots and do not grow deep.

9.77 Roots absorb water, Tradescantia
See diagram 9.77: Roots absorb water
Put a rooted and an unrooted shoot of Tradescantia each in a test-tube. Select the shoots so that they have the same leaf area. Fill two test-tubes with tap water to 2 cm below the rim. Pour a thin layer of paraffin oil on the water in each test-tube. Mark the height of the surface of the liquid on the test-tube with a felt pen. Put the test-tubes in a test-tube rack. Record the level of water in the two test-tubes each day. The surface of the liquid drops slightly in the test-tube containing the unrooted Tradescantia shoot, but drops more in the other test-tube containing the well-rooted shoot.

9.78 Celery stalk, bean stem
See diagram 9.78: T.S. Celery stalk | See diagram 9.3.60: T.S. Bean stem
Celery stalks are enlarged petioles. A petiole is not a stem. The stem of the celery plant is reduced to a disc.
Experiment
Wear safety glasses and nitrile chemical-resistant gloves. Use a one-sided razor blade to cut thin transverse sections and longitudinal sections from a celery stalk. Be careful! Cut away from the body. Mount in water and apply a coverslip. Stain sections with acid phloroglucin.
Observe the following tissues:
1. Epidermis is a single layer of cells with a thick cuticle covering the outermost surface.
2. Collenchyma has cellulose thickenings in the corners of the cells.
3. Parenchyma has large cells with thin cell walls.
4. Vascular tissue is arranged as discrete collateral bundles. Each vascular bundle has phloem to the outside, xylem to the inside and cambium between.
5. Sclerenchyma caps the vascular bundle.

9.79 Dicotyledon stem, sunflower
See diagram 9.79: T.S. Young sunflower stem | See diagram 9.78.1: Sunflower Stem LS
1. Mount the section and irrigate with an aniline salt, to stain the woody tissues. Examine the whole section under low power. Observe the epidermis, cortex and central cylinder. Look for any layers of the cortex thickened to give additional strength. Observe the vascular bundles composed of xylem, phloem and cambium. Observe the pith and note if the stem is solid or hollow.
2. Observe the stem under high power and note the cellular structure in detail. Observe the epidermal tissues, cortical tissues and a complete vascular bundle.
3. Examine a similar stem by means of radial longitudinal sections, R.L.S. Note the appearance of the collenchyma, the annular protoxylem vessels, and the pitted metaxylem vessels.

9.80 Monocotyledon stem, maize, (corn), Zea mays
See diagram 9.80: T.S. stem of maize (corn)
Use low power to observe the small size, large number, and irregular arrangement of the vascular bundles. The outer scattered vascular bundles are surrounded by fibres to strengthen the stem. Note the complete absence of cambium.

9.80.1 Monocotyledon stems, Cocos nucifera coconut, Dracaena
See diagram 9.78.4: Dracaena | See diagram 53.4: Cocos nucifera, coconut

9.82 Leaf of Hakea, xeromorphic leaf
See diagram 9.69.1: Hakea stomate
Many Hakea species have needle-shaped leaves in which there is a reduction in leaf surface / volume ratio.
Observe the following:
1. epidermis with thick cuticle and sunken stomates,
2. palisade mesophyll cells in a double layer interspersed with sclereids,
3. central storage mesophyll with scattered vascular bundles.

9.97 Roots from plant parts
See diagram 9.85: Potato
Obtain a box of sand and put it out of direct sunlight. Wet the sand thoroughly and keep it moist.
Plant any of the following in the sand:
1. various bulbs,
2. cuttings of begonia and geranium stems,
3. a section of sugar cane stem with a joint buried in the sand,
4. a section of bamboo stem with a, joint buried in the sand,
5. carrot, radish and beet tops, each with a small piece of root attached,
6. an onion,
7. an iris stem,
8. pieces of potato containing "eyes",
9. a branch of willow.

Table 9.0.1 Characteristics and functions of plant tissue types.
Tissue system Tissue types (numbered) Characteristics Function
Meristematic tissues 1. Apical meristems
2. Cambium
Closely packed cells, large nuclei, thin walls Produce new cells by cell division
Ground tissues 3. Parenchyma: Unspecialized ground tissue especially cortex and pith Living protoplasts, cells loosely-packed, thin cellulose walls, simple pits
3a Chlorenchyma has chloroplasts
3b Aerenchyma has large intercellular spaces
Packing tissue, lateral transport, mechanical support by turgidity in herbaceous plants, cells can divide after wounding and produce cambium
Ground tissues
4. Collenchyma
Subepidermal or in cortex, only in stems and leaves
Living protoplasts, elongated cells, primary cellulose walls, thickened corners, simple pits Supporting tissue in strands or cylinders, subepidermal in stems, petioles and leaf veins
Ground tissues
5. Sclerenchyma: Fibres in stem cortex and leaf mesophyll tissue and sclereids scattered in parenchyma Cells with thick lignified walls, dead at maturity 7 Fibres are interlocking elongated and narrow cells 8 Sclereids have variable size and shape Strengthening tissue of root, stem
and leaf Fibres in strands or cylinders in cortex Small groups of sclereids in parenchyma of soft leaves and fruits, or massed to form stony tissue of fruits
Dermal
tissues
6. Epidermis: Layer covers primary body of plant One cell in thick with cutin on outer wall Stomata (pores) on aerial plant parts Protective and prevent desiccation in
stem and leaf Stomata allow gas exchange Epidermal root hairs increase water uptake
Dermal
tissues
7. Periderm: External covering replaces epidermis in secondary body Phellem (cork): Layers of cells produced outside by cork cambium (phellogen) protoplast dead at maturity, walls impregnated with suberin Protective waterproof layer replaces epidermis in older stems and roots Lenticels on stems have function of stomata
Vascular
tissues
8. Xylem: Consists of vessels, tracheids, fibres and parenchyma cells 8a Vessels: Tubes formed by dissolution
of the end walls of a vertical column of cells Walls have lignin and bordered pits, except in protoxylem, no protoplast at maturity
Water conduction through long continuous tubes Protoxylem annular
and spiral thickenings allow extension of vessels (and tracheids) in regions of elongation
Vascular
tissues
Xylem
8b Tracheids: Elongated lignified cells have complete end walls, have bordered pits except in protoxylem, no protoplast at maturity Water conduction through pits in walls, or through non-lignified areas of walls as in protoxylem spiral and annular tracheids
Vascular
tissues
Xylem
8c Xylem fibres: Like sclerenchyma fibres Strengthening but not conducting
Vascular
tissues
Xylem
8d Xylem parenchyma: In vertical
columns, may have lignified walls
Food storage

Vascular
tissues
Xylem
8e Xylem ray parenchyma: From ray initials of cambium, radially elongated Radial conduction of food and water across xylem
Vascular
tissues
9. Phloem: Consists of sieve tubes, companion cells, fibres, parenchyma and ray parenchyma 9a Sieve tubes: Vertical rows of elongated cells have perforated end walls (sieve plates), have cytoplasm but no nucleus at maturity Conduction of organic food materials


Vascular
tissues
Phloem
9b Companion cells: Elongated cell has nucleus and dense cytoplasm, with sieve tube element, many mitochondria Control of sieve tube



Vascular
tissues
Phloem
9c Phloem parenchyma: Vertical files of parenchyma Storage of foods, tannins and resins
Vascular
tissues
Phloem
9d Phloem fibres: Like fibres of sclerenchyma Strengthening
Vascular
tissues
Phloem
10 Ray parenchyma: From ray initials of cambium, radially elongated Radial conduction across phloem