Plant Anatomy: A Practical Guide - Paniuta O.O. 2019

Topic 1. The Cell
Laboratory Work No. 1. Structural Features of the Plant Cell under a Light Microscope

Theoretical Background. Living plant Cells consist of a Cell wall and internal contents. The Cell wall encloses the cytoplasmic contents on all sides, serving as a protective Structure.

In young cells, the wall is very thin, delicate, elastic, and transparent. It is composed of Cellulose, or dietary fiber—a substance belonging to the polysaccharide carbohydrate group with the general formula (С₆Н₁₀О₃)_n. The cell wall also contains a certain amount of pectic substances, which are similar in COMPOSITION AND PROPERTIES to CARBOHYDRATES. Pectic substances fill the spaces between the cellulose fibrils, forming the intercellular substance that cements together the walls of adjacent cells.

The properties and COMPOSITION OF THE cell wall change as the cell and the entire plant develop. It becomes thicker and develops various types of uniform or non-uniform thickenings. In some areas, thin spots without thickenings remain, which are known as pits. Thickenings vary in Morphology and can form rings, spirals, reticulate networks, etc. In mature cells of many plants, The chemical composition of the wall also changes: it may undergo lignification, suberization, mucilaginous degeneration, cutinization, mineralization, etc.

If the intercellular substance breaks down, the cells dissociate and plant Tissues disintegrate into individual cells. This phenomenon is observed during tissue maceration.

Cell shape varies even within the same plant. Two primary cell shapes are distinguished: parenchymatous and prosenchymatous. Parenchymatous cells may be spherical, polygonal, plate-like, stellate, etc. Prosenchymatous cells are elongated, sometimes reaching 1 cm or more in length, and are referred to as fibers.

Cells are generally small, and their dimensions vary among different plants. The diameter of elongated cells does not exceed a few micrometers (thousandths of a millimeter). In fiber crops, cells are significantly larger, exceeding 1 cm in length. For the most part, plant cells are so small that they cannot be seen with the naked eye.

Beneath the cell wall lies the protoplast, which primarily consists of the Cytoplasm, Nucleus, Plastids, and other Organelles. The Nature of the cell contents changes throughout the cell's ontogeny. For instance, young cells contain cytoplasm, a nucleus, Mitochondria, and plastids. Later, as the cell develops, one or more vacuoles appear. Plastids also undergo changes: some remain colorless (leucoplasts), while others transform into green plastids (METABOLISM/14.html">Chloroplasts) and yellow ones (chromoplasts). Subsequently, other structures emerge within the cells. As a result of Photosynthesis, starch and oil develop in the cytoplasm, while soluble sugars and Other Compounds accumulate in the cell sap.

In young cells, the cytoplasm fills the entire cavity, whereas in older cells, it is sparse, forming a thin parietal layer while the rest of the cavity is filled with cell sap, occasionally traversed by cytoplasmic strands extending from the parietal layer. Cell sap is mostly colorless, though in some plants, it is pigmented red, blue, or violet. These pigments are collectively known as anthocyanins. The color of the cell sap also depends on the pH of the medium: an acidic reaction results in a red color, while an alkaline reaction yields a blue color. When a cell dies, its internal contents disappear, leaving only the cell wall. Such dead cells are abundant in old perennial trees, where they perform crucial mechanical and protective Functions, imparting structural strength and shielding the plant from Water evaporation and damage.

To examine living plant cells under a Microscope, one must prepare thin sections to create microscopic slides. Many specimens can be observed whole in a drop of water, such as Elodea canadensis and Elodea densa, or a strip of onion scale epidermis where cells are arranged in a single layer.

Elodea canadensis is commonly found in nature in stagnant water bodies and is easily cultivated in aquaria, whereas Elodea densa is grown exclusively in greenhouses and warm aquaria. To prepare a slide, a leaf is detached from the Elodea stem using tweezers and examined in a drop of water under a microscope. The leaf consists of two cell layers, with the upper layer containing larger cells than the lower one. For examination, it is best to use leaves from the apical, actively budding region of the plant.

The green coloration of the leaf cells is not uniform, which is due to varying concentrations of chlorophyll within the cells.

Living cells can also be observed under a microscope using onion ROOT tips and root hairs.

Young root tips developing root hairs are best suited for this purpose. Such tips are cut with scissors, transferred to a drop of water on a microscope slide, and covered with a coverslip.

To examine the living cells of aerial plant parts, trichomes (hairs) are ideal; they can be excised from the stems and petioles of cucurbits, stinging nettles, apple trees, and other plants. An exceptional subject for such studies is the staminal Hair of Tradescantia virginiana. Slides from this plant are prepared as follows: holding a flower bud from a young inflorescence in the left hand between the thumb and forefinger with the Base of the flower pointing upward, a cut is made near the base. This cut passes through the attachment point of the sepals, petals, and stamens. In a drop of water, the anthers are removed, and the staminal hairs are covered with a coverslip. Microscopic examination reveals that they consist of elongated, barrel-shaped cells.

Living cells from the pulp of watermelon, tomato, and other plants are also clearly visible under a microscope.

Objective: To study the Structural Features of a living plant cell.

Materials and Equipment: Light microscopes, Glass slides and coverslips, dissecting needles, tweezers, glass stirring rods, filter paper, scissors, razor blades, distilled water, iodine-potassium iodide solution, aniline green, chloral iodine, plant material.

Slide: Structure of Epidermal Cells in Garden Onion Scales (Allium cepa L.)

This specimen is exceptionally well-suited for studying cellular structure.

The main advantage of this specimen is that cellular morphology can be investigated using fresh, living material at any time of the year.

When studying The structure of onion scale epidermis, four separate slide preparations are made, each treated with different stains to highlight fine cellular details. To prepare the specimen, select a pigmented (purplish-red/blue-tinged) onion bulb. Using a scalpel, peel off one colored scale. Holding it in the left hand with the convex side facing up, use the tip of a sharp needle in your right hand to puncture the edge, grip it with tweezers, and peel it upward and sideways. The gripped area may not be extremely thin, as it might include underlying tissues, but the edge of the epidermis will typically appear as a thin, nearly transparent film. Place this film into a drop of water, using a razor blade to trim off any thick edge that would prevent the coverslip from lying flat on the glass slide.

When placing the film into the drop of water, ensure it faces outer-side up and is thoroughly flattened to avoid air bubbles between the coverslip and the slide. To remove air bubbles, lift the coverslip, add another drop of water, Touch one edge of the coverslip to the slide, and gently lower it. If water spills out from under the coverslip, carefully blot it away with filter paper and ensure the top surface of the coverslip is completely dry. Examine the prepared slide first under low magnification to locate the thinnest and clearest region, and then switch to high magnification.

The epidermal cells of onion scales are elongated and fit tightly against one another.

Under high-magnification observation of a thin preparation, it is clearly visible that the lateral cell walls are not continuous, but appear intersected by small, delicate regions known as pits.

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Fig. 2. Epidermal cells of an onion (Allium cepa) scale leaf:

1 - cell wall, 2 - cytoplasm, 3 - nucleus, 4 - vacuole

The walls of the scale epidermal cells are colorless and transparent. To examine both the upper and lower cell walls, use the fine adjustment knob to focus the objective for the best view; however, this will make the cell boundaries and internal contents appear very faint. For a more detailed examination of the cell contents, observe the cell in optical cross-section by adjusting the microscope tube until the lateral walls come into sharp focus.

Upon examining the slide (Fig. 2), a thin layer of granular cytoplasm can be seen lining the cell wall. At the cell poles and around The Nucleus, the cytoplasm gathers into denser masses from which cytoplasmic strands extend across the cell to merge with the parietal layer.

The nucleus is clearly visible within the cytoplasm, located either centrally or near the lateral wall, but always embedded in the cytoplasm. The nucleus contains one or two nucleoli.

The cell lumen is occupied by vacuoles filled with cell sap, which is colored violet by anthocyanin.

After careful examination of the slide, staining can be initiated. To do this, place a strip of filter paper against one edge of the coverslip while adding 2-3 drops of aniline green solution to the opposite edge. Aniline green intensely stains the nuclei. A diluted solution should be used, as this dye also stains the cytoplasm and cell walls.

Once the slide has been sketched, remove it from the stage, take a second slide, and treat it similarly with an iodine-potassium iodide solution. This turns the cytoplasm yellow and imparts a yellowish-brown color to the nucleus.

A third slide is treated with chlor-zinc-iodine. Since the upper and lower walls lie in the plane of the glass and stain blue, the entire preparation will appear blue. By adjusting the focal length with the fine adjustment knob, the cell contents—the cytoplasm and nucleus—can be observed stained in yellowish-brown.

In the preparation treated with chlor-zinc-iodine, unthickened areas of the cell wall—pits—stand out distinctly.

Slide. Leaf cells of Canadian waterweed (Elodea canadensis Michx.)

Place an Elodea leaf on a microscope slide in a drop of water so that its morphologically upper surface faces upward, and gently cover it with a coverslip, taking care to avoid air bubbles.

Examine the prepared slide under low magnification. The leaf blade consists of two cell layers, with the upper layer cells larger than the lower ones. Intercellular spaces filled with water or air are visible between the cells of both layers. The Cells of the leaf blade have a parenchymatous structure. A distinct midrib, consisting of several layers of elongated cells with green contents, is clearly visible in the center of the leaf blade. Elongated cells are visible along the margins of the leaf blade, some of which form small Teeth. Under high magnification, a thin cell wall and cytoplasm containing the nucleus and chloroplasts can be observed. The cytoplasm is located adjacent to the cell wall. In the center of the cell is a vacuole filled with colorless cell sap.

Slide. Leaf hairs of spiderwort (Tradescantia virginiana L.)

Spiderwort leaf hairs are an exceptionally convenient material for studying Cell Structure.

To prepare the slide, use scissors or a scalpel to cut a thin strip from the base of the leaf—where it presses closely against the stem and is densely covered with hairs—place it on a microscope slide in a drop of water, and cover with a coverslip. Lower the coverslip from the base of the hair toward its apex to expel any trapped air underneath.

Examining the slide under low magnification reveals that the hairs are arranged in neat rows along the leaf margins. Each hair consists of cells that are short and wide at the base and long and narrow at the tip. The terminal cell is short and pointed at the apex. Switching the microscope to high magnification shows that the cell walls are thin and double-contoured. The cytoplasm lining the wall surrounds the nucleus and extends across the cell via cytoplasmic strands, forming a reticular network. The rounded nucleus is clearly visible under the microscope—it is large and denser than the rest of the cell contents. The cytoplasm also contains leucoplasts, which appear as small, highly refractive white granules.

Slide. Cell structure of watermelon pulp (Citrullus vulgaris Schrad.)

To study living Cells under the microscope, isolating individual cells from watermelon pulp is most convenient. Take a piece of pulp from a ripe red watermelon, where individual bubble-like cells are clearly visible due to the dissolution of intercellular substance. Separate them and use a brush or dissecting needle to transfer them to a microscope slide, adding a drop of the juice released from the watermelon.

Fig. 3. Watermelon pulp cells:

1 - cell wall, 2 - cytoplasmic streams, 3 - nucleus with nucleolus, 4 - vacuole, 5 - chromoplasts

Mix everything gently on the microscope slide and examine under low and high magnification. Individual clear cells are visible under the microscope (Fig. 3).

When examining this slide, the Structural components of the cell can be observed, namely: the colorless, transparent cell wall and the internal contents consisting of granular grayish cytoplasm, orange-red plastids (chromoplasts), a nucleus with a nucleolus, and cell sap.

After examining the slide under both low and high magnification, draw a careful sketch of it in your laboratory notebook.

Slide. Structure of a tomato flesh cell (Lycopersicon esculentum Mill.)

Cell structure can be studied using slides made from a red tomato.

To do this, use a sharp scalpel to remove a small piece of flesh along with the epidermis, place it in a drop of water on a microscope slide, and tease it apart with a dissecting needle to spread the mass into an even layer. Cover the prepared slide with a coverslip and examine it first under low magnification, and then under high magnification.

Tomato flesh cells vary in size and shape: round, oval, or elongated (Fig. 4).

Fig. 4. Tomato flesh cells:

A - cells of ripe tomato flesh in their natural state, B - cells after Treatment with chlor-zinc-iodine; 1 - cell wall, 2 - cytoplasm, 3 - chromoplasts, 4 - starch grains, 5 - nucleus, 6 - vacuoles

The illustration shows thin cell walls lined on the inside with a narrow layer of granular cytoplasm. Cytoplasmic strands extend from this layer in various directions. Visible within the cytoplasm are chromoplasts and starch grains, as well as the nucleus, which is located either in the center of the cell or near its wall. The nucleus is round or oval in shape. The cytoplasmic strands form a network within the cell enclosing vacuoles of various sizes, which appear as irregular orange-colored plates. The riper the tomato, the more chromoplasts it contains. The cell sap inside the vacuoles is colorless. To examine the slide thoroughly and sketch it in your lab notebook, you need to treat it with various Reagents. To do this, place a strip of filter paper on one side of the coverslip and apply 1–2 drops of an iodine-potassium iodide solution to the other side.

Under the action of iodine, plasmolysis begins in the cell, causing the cytoplasm to pull away from the cell walls. Iodine stains the cytoplasm yellow, the nucleus brown, and the starch grains blue. After careful examination, compare the slide with the illustration in your lab manual to identify details that were less visible in the unstained preparation.

Prepare another slide on a separate microscope slide and treat it with chlor-zinc-iodine, taking care not to use an excess of the reagent. Cover the finished preparation with a coverslip and examine it under a microscope. Treated with chlor-zinc-iodine, the cytoplasm and nucleus stain brown, the cell wall stains blue-purple (this is the characteristic reaction of chlor-zinc-iodine with cellulose), and the starch grains stain black-blue. The cell wall stains unevenly, making the thicker areas appear darker.

To make the nucleus stand out more clearly on the slide, treat it with stains that intensely color only the nucleus while leaving the cell wall, cytoplasm, and other cell components unstained. Such stains include aniline green, methyl green, and carmine.

When staining slides with aniline green, place a small piece of tomato flesh on a microscope slide in a few drops of a weak solution of this stain and thoroughly tease it apart with a dissecting needle to separate individual cells. Then, cover the slide with a coverslip and let it stand for a few minutes. During this time, the dye penetrates the cell wall, reaches the nucleus, and stains it bright green. One or two nucleoli are visible within the stained nucleus.

Note. Very weak dye solutions should be used for staining slides, because a strong solution will stain not only the nucleus, but also the cell wall and cytoplasm. The same thing happens if the staining process takes too long.

In addition to the aforementioned specimens, the structure of a plant cell can also be examined using: pumpkin leaf trichomes, onion scale epidermis, fuchsia petal epidermis, moss leaves, and Elodea leaves.

In most of the specimens listed above, the following cell components can be identified: the cell wall, cytoplasm, and nucleus. Cytoplasmic strands extend from the parietal layer of cytoplasm toward the center of the cell, clustering densely around the nucleus. The nucleus may be spherical or oval. A nucleolus is often visible within the nucleus. Vacuoles are filled with cell sap. In the cytoplasmic cells of watermelon flesh, fuchsia petals, and other plants, chromoplasts can also be observed, appearing as orange-red granules. Lab drawings should depict the following cell parts: cell wall, cytoplasm, nucleus, nucleolus, vacuoles, and plastids.

Slide. Structure of a pumpkin trichome cell

(Cucurbita реро L.)

Pumpkin trichomes ( hairs) serve as an exceptionally convenient specimen for studying cell structure. To make a slide, it is best to use trichomes from fresh, green pumpkin leaves. Place a drop of water on a microscope slide and gently scrape trichomes from the lower surface of the leaf into the water using a scalpel. Next, cover the preparation with a coverslip and examine it first under low and then under high magnification, having selected the clearest area of the slide.

Fig. 5. Pumpkin trichome cells:

1 - cell wall, 2 - cytoplasm, 3 - nucleus

The cell contents are enclosed by a thin cell wall (Fig. 5); inside the cell is the cytoplasm, which forms a thin layer tightly pressed against the wall. Cytoplasmic strands extend from this parietal layer, crossing the cell in all directions. A relatively large and dense Cell Nucleus, darker than the cytoplasm and therefore very clearly visible, lies within the cytoplasm. The cytoplasm surrounding the nucleus has a distinct granular structure. The entire volume of the cell in the spaces between the cytoplasmic strands is filled with cell sap. Chloroplasts are also clearly visible in living cells from fresh material. Living pumpkin trichomes provide an excellent specimen for observing cytoplasmic streaming (cyclosis).

Note. Pumpkin trichomes are convenient for observation because they can be examined in their normal, living state. It is recommended to use this material for practical work whenever live green leaves are available on pumpkin plants.

Slide. Moss leaf cells (Plagiomnium cuspidatum (Hedw.) T.J. Kop.)

To study moss leaf cells, take a small piece of a moss stem and use tweezers to pluck off a leaf. Rinse the leaf in water, place it on a microscope slide in a drop of water, and cover it with a coverslip. A moss leaf does not fit entirely within the microscope's field of view; therefore, to get a clear picture of its shape, structure, and components, the slide must be moved around.

The moss leaf is plate-shaped and consists of a single layer of polygonal parenchymatous cells. Along the edges of the leaf blade, distinct teeth are visible, formed by elongated prosenchymatous cells.

In the parenchymatous cells, a polygonal wall and living cytoplasmic contents are clearly visible. The cell walls consist of three layers: two thicker layers belonging individually to each cell, and a thin layer located between them. This forms the so-called middle lamella, which is composed of pectic substances and plays a crucial role by providing strong adhesion between adjacent cells.

The cell contents consist of cytoplasm and chloroplasts. The chloroplasts vary in shape, being oval, elongated, etc. In the light, colorless starch granules can be observed within the chloroplasts. The nucleus is difficult to spot due to the high density of chloroplasts.

The Functions of the various cells in the moss leaf blade differ. Photosynthesis takes place in the parenchymatous cells containing chloroplasts, whereas the marginal prosenchymatous cells that form the teeth perform a mechanical function—they reinforce the edges of the leaf blade.

In the middle of the leaf blade, the so-called leaf midrib is visible. It consists of several layers of elongated cells.

Water and dissolved nutrients are transported through the cells of the midrib.

Slide. Cells of fuchsia flower petals (Fuchsia hybrida hort. ex Siebert & Voss)

Fuchsia flower petals serve as an exceptionally convenient subject for studying plant cells. To prepare the slide, a single petal is taken from a fuchsia flower, and a thin layer of epidermis is peeled off using forceps. This is placed on a microscope slide in a drop of water, covered with a coverslip, and examined under a microscope at low and high magnifications. Microscopic observation clearly reveals that the epidermis is composed of elongated cells that fit tightly against one another. The cell walls are very delicate and thin, displaying a wavy rather than straight contour. The cells contain fine-grained cytoplasm, a large nucleus with one or two nucleoli, and—in some cells—vacuoles that are either colorless or pigmented red by anthocyanin.