To the Fun Science Gallery Contents

 

4 - Flowering Plants (Phanerogams)

Giorgio Carboni, April 2007
Translated by Sarah Pogue

 


 

CONTENTS
 
INTRODUCTION
ORGANS AND TISSUES
Flowers
Fruits and seeds
Method of seed dispersal
Roots
Stems
Meristems
Leaves
Epidermal and hairy tissues
WET MOUNTS
FOR THE LITTLEST ONES
IN DEPTH
CONCLUSION
BIBLIOGRAPHY
INTERNET RESOURCES

Figure 1 – Ovule of white campion flower (Silene latifolia).
Detail of figure 14. Ovule dimension = 1 mm.


INTRODUCTION

In the previous article, we dealt with the “inferior” terrestrial plants, those plants which reproduce by means of spores and are called Cryptogams. In the present article, we will look at the “superior” terrestrial plants, or more exactly, the Phanerogams. These are vascular plants that have flowers and produce seeds. The terms Spermatophyte (from the Greek: spčrmatos = seed; phytňn = plant), Phanerogam (fanerňs = evident; gŕmos = marriage) and Anthophyte (from ŕntos = flower) are synonymous. These include the Gymnosperms (plants with flowers and unenclosed ovules: conifers and other minor families) and the Angiosperms (plants with flowers and closed ovaries: the most common plants).

In contrast to the Cryptogams, which are relatively primitive plants, the Phanerogams and in particular the Angiosperms are plants that have undergone a notable evolution and which have a wide variety of specialised tissues and organs such as roots, stems, leaves, flowers etc.


ORGANS AND TISSUES

FLOWERS


The flowers are without doubt the most striking and beautiful parts of the plant, and it is with these that we will begin our explorations. Now, think of a flying insect that catches sight of a flower, changes direction, lowers its altitude and as it approaches detects an intoxicating perfume as the petals seem to open up to reveal a landing strip. The insect lands and goes towards a perfumed coloured chamber where it finds nectar and pollen. The encounter with the flower will be a fascinating experience also for us. Unfortunately, we cannot taste the nectar, but the colours and the shapes that we can admire are certainly delightful, particularly if we observe them under the microscope.

During the spring, in fields and gardens, you can collect an infinite number of flowers to examine. You mustn’t think that when observing a flower under the microscope you can see the same things as when you look at it with the naked eye only bigger, because flowers almost always reveal unexpected details when viewed with a microscope which are very beautiful and of great interest.

Take for example the heartsease or wild pansy (Viola tricolor, Figure 2). You will have noticed how its petals are velvety. With the stereoscopic microscope, it is possible to note that the petals are covered with finger-like cells called papillae (Figure 3). The papillae are what makes the petals of the pansy particularly velvety to the touch. The height of these “fingers” grows as you move towards the interior of the flower and when you are close to the gynoecium or pistil these suddenly reach a length of several millimetres and are called hairs (Figure 4). Thanks to these finger-like structures and their colours, the observation of the pansies is particularly striking. There are different species of pansy and it is certainly interesting to observe the differences between them.

 

Figure 2 – Flower of heartsease/wild pansy.

Figure 3 - Papillae from the upper surface of the petal.
Passo = Distance between papillae = 0.02 mm.

Figure 4 – Hairs close to the pistil.

Figure 5 – Hairs of the pansy viewed under the
biological microscope. Diameter = 0.14 mm

Figure 6 – Nuclei of pansy hairs.

Figure 7 – Cells from the lower surface of the petal.

 

With a pair of fine tweezers, remove a piece of tissue from a petal where the “fingers” are longest and observe it under the biological microscope (Figure 5). Obviously, in order to do this, you must immerse the strip in water, then place it between two microscope slides and add some drops of alcohol to disperse any air bubbles present. You will realise that these hairs are none other than single cells and you will also see their nucleus (Figure 6). The papillae that cover the petal and that give it its velvety aspect and texture are cells and on every petal there are tens of thousands of these (Figure 3). Obviously, each cell has its nucleus and this is normal, but it makes us reflect on the complexity and perfection of biological structures. If necessary, colour the strip with a dye for chromatin in order to make the nuclei more evident.

Remove the strip of the epidermis from the underside of the petal and look at it under the microscope. You will see that the single cells of this tissue have a shape that reminds us of jigsaw pieces (Figures 7 and 41). Often the flowers, but also the leaves, possess cells of this shape which allow the plant to increase the surface area of contact between the cells.

 

Figure 8 – Diagram of a flower.

Figure 9 - Campanula flower (Campanula sp.).

 

Now we will move on to a more detailed examination of the flowers in which we will seek to understand their form and organisation. We will begin with the examination of simple flowers in which it is easy to distinguish the four principal parts: sepals, petals, stamens and pistil (Figure 8). Flowers suitable for this examination are the campanula (Figure 9), ranunculus, dianthus and many others. Many other flowers, amongst which is the violet, are not well-suited for this initial observation because their organs are highly modified with repect to the “basic” structure and it would be difficult to recognise them. Further on, when you are familiar with the wide variety of botanical structures, you can also examine these flowers.

Now, arm yourself with tweezers and a scalpel and, under the stereoscopic microscope, section the flower to analyse its parts:
- remove a couple of petals to uncover the pistil and stamens;
- remove a stamen and observe the anther and the pollen that it contains;
- examine the various parts of the pistil: ovary, style, stigma. In the campanula the stigma is divided into three parts;
- look and see if there are pollen grains on the stigma;
- section the pistil longitudinally, examine the ovary, the placement of the ovules and their stage of maturation;
- observe the contour of the petals and their upper and lower surfaces, describe any hairs present and any other characteristics;
- observe the sepals, especially any hairs that cover them;
- describe any insects present on the flower.

With a biological microscope:
- observe the pollen at a high magnification;
- remove a strip from the upper and lower surfaces of the petal to observe the cells;
- make transverse sections of the petals;
- observe any hairs present on the petals and sepals.

During this anatomical exam, it would also be useful to make a drawing or a diagram of the sectioned flower and to write a brief report accompanied by drawings and photos. At the end you could collect all the drawings, photos, reports and preserved specimens in a file. These documents will be very useful for comparative examinations of the flowers of different species.

After studying the flowers of different plants, you will realise that the differences regard not only the colour, but also and above all the structure. In fact, during the course of evolution, the “basic” form of the primitive flowers has greatly diversified and the study of these is now particularly interesting. For example, think of the daisy which is formed of hundreds of miniscule flowers, each of which is equipped with stamens and pistil. These flowers are arranged in spirals on the flower head. It is not by chance that these flowers are called Composites. The flower head is the structure composed of all the small flowers. With regard to the terminology, it is incorrect to call the flower head of a Composite a flower and instead this must be called an inflorescence or a flower head or better a capitulum.

 

Figure 10 – Chicory inflorescence
(Cichorium intybus).

Figure 11 – Chicory floscule: note the ovary low
down, the stigma, the blue anthers and the petal.

Figure 12 – Stigma of a chicory inflorescence.
Numerous white pollen grains are visible.

 

Chicory is also a composite, even though the number of flowers of which the inflorescence is composed is inferior to that of the daisy. In summer, chicory is very common in fields and along the roadside. Its azure inflorescences are highly visible (Figure 10). As with other composites, the petal edges are serrated. Sectioning the inflorescence of a chicory plant, you can ascertain that each of its small flowers has a corolla consisting of a single petal. This little flower is called a floscule (Figure 11). At its base is the ovary which contains a single ovule that will become an achene (a dry fruit containing a single seed). The anther is formed from the fusion of 5 elements, is tubular, blue in colour and wrapped around the pistil. From the anther the Aries-shaped stigma protrudes, upon which numerous white pollen grains are visible (Figure 12). While you look at the interior of the inflorescence you may see some tiny insects peeping out from amongst the floscules; these will most likely be thrips, whose feathery wings you should also observe.

Observing the inflorescence of chicory, this humble and common plant, and admiring the spectacle offered by its corolla, you will realise how nature is full of wonders that don’t cost anything. All you need is a stereoscopic microscope, at times even just a magnifying lens, to admire the splendid forms that nature tirelessly produces. This is how a flowering field which, to an ordinary person doesn’t offer anything in particular, to a microscopist appears instead as a source of infinite wonder.

Another interesting flower is the white campion (Silene latifolia), a plant that flowers between May and November. As you can see in the figures below, the calyx of this flower has the shape of a flask from which a crown of white petals emerges. This species is dioecious, that is, the male and female reproductive organs are on separate plants so that on one plant you will find flowers with the pistil and on another flowers with the stamens. The male flowers have a narrow calyx and from this the anthers emerge, the female flowers have a swollen calyx due to the presence of the ovary and from this the stigmas emerge (Figures 13, 14 and 15).

 

Figure 13 – Female white campion flower (Silene
latifolia
). During maturation, the ovary swells.

Figure 14 – White campion ovary (with calyx) sectioned.
Diameter approximately 16 mm. See details in figure 1.

Figure 15 – Male white campion flower. On the right, the
open flower. Note the stamens and the absence of the ovary.

 

- Collect the female flowers of the white campion at different stages of maturity;
- take a female flower that still has petals (Figure 13);
- with a razor blade, remove the calyx and expose the ovary. Note that the petals depart from the base of the ovary and note also the absence of stamens;
- section the swollen ovary longitudinally and observe the ovules (Figure 1 and 14);
- take a male flower. With a small pair of scissors, open the calyx and if necessary remove a couple of petals;
- observe the stamens, the anthers and the pollen. Note the absence of the ovary.

After the “basic” flower such as that depicted in Figure 8, I presented the flower of the campanula, one of the composites and the flower of a dioecious plant, in order to prepare you for the diversity of flower structure before leaving you to continue your observations alone.

 

EXPERIMENTS ON POLLEN GERMINATION

The flower stigma is often sticky due to the presence of sugary substances. Inserting pollen grains in a sugary solution, it is possible to provoke the emission of the pollen tube. The concentration of sugar in this solution should be between 2 and 20%, according to the species from which the pollen derives. In the majority of cases, a concentration of 10% is fine, but for the Composites it should be from 30 to 45%. Boil some water with sugar added and prepare some microscope slides for Van Tieghem observations. Another observation method is that of inserting the pollen grains and solution between normal microscope slides. If the grains are big, it will be necessary to insert some pieces of broken coverslip between the slide and the coverslip, in such a way as to avoid squashing them. Every so often it will be necessary to add a little water to the solution as it evaporates. Some texts suggest adding a little gelatine to the solution (approximately 2%). After boiling the solution to sterilise and homogenise it, it will be necessary to keep it in a bain-marie at 25°C to keep it fluid, then place a thin film of fluid on some clean sterile microscope slides. On this film place some pollen grains, even of different species, and keep them in a humid environment, such as in a Petri dish or in a small shallow glass jar containing a strip of paper soaked in water. The emission of the pollen tube can take many hours and will not necessarily happen. In the case that the emission does take place, you could try to highlight the three nuclei that are emitted in the tube with a chromatin colourant such as Toluidine blue or Methylene blue. It is possible to experiment with different concentrations of the sugary solution to adapt it to different plant species. The pollen of the plant commonly known as "busy lizzy" (Impatiens sultani or Impatiens walleriana), emits tubes in less than an hour in a 10% glucose solution. In the interior of the pollen tube it is also possible to observe the cytoplasmic currents.

Figure 16 – Grain of pollen from the lesser celandine
(Ranunculus ficaria) which has produced a pollen tube.
Kept in a 15% sugar solution for approximately 6 hours.
Phase contrast. Field = 0.25 mm.


FRUITS AND SEEDS

The flower is a specialised structure of the Spermatophytes dedicated to sexual reproduction. The fertilised flower is destined to undergo a transformation which generally leads to the formation of fruit and seeds. The fruit develops from the lower wall of the pistil (ovary) while the seed develops from the fertilised ovule.

That which we consider to be the fruit and which represents the edible part of, for example, an apple, in botany is not considered as the true fruit since it develops from the receptacle (tissue found at the base of the flower). In botany, the structure derived from the ovary is considered to be the fruit, which protects the seeds and which in an apple corresponds to the core.

Often, during maturation, the ovary and the style transform significantly. Section the ovary at different stages of maturity and try to follow the development of the fruit, observe the development of the seeds and the mechanism for their dispersal. Look at, for example, a pumpkin cut in two. In the lower part you will see what remains of the flower, in the upper part the petiole of the pumpkin which at one time supported the flower, and in the central cavity the ovules by now transformed into seeds. All the rest of the pumpkin is a transformation of the ovary. Analysing the seeds, it will be possible to distinguish the embryo with its cotyledons, endosperm, reserve tissue that surrounds and provides nutrition for the embryo, and the external integument. The text indicated in [404] is a guide to the spontaneous flowers and is very well illustrated. Besides the entire plants, it shows details regarding the flowers and fruits which can aid you in understanding their structure.

The carpel is a leaf that during the evolution of the Angiosperms transformed into the ovary. The pistil can be formed from one or more carpels. Fruit can be simple (e.g. peach), aggregate (e.g. raspberry) or multiple (e.g. pineapple). The fruit can be fleshy or dry. Do not expect, therefore, for the fruit to always be similar to an apple, as it could also be a pod as in the case of the bean, a winged achene like the maple etc. Examine and describe the fruits and seeds of different plants (e.g. Figures 17, 18, 19, 20, 21, 24 and 42). In doing this, it will be useful to you to familiarise yourself with the botanical classification of fruits and the terminology which you can find in a biology textbook [001] or in a botanical atlas [003] and [004].

 

Figure 17 – Oat seeds.

Figure 18 – Shepherd’s purse ovary opened to show
the ovules. The ovary is approximately 4 mm in width.

Figure 19 – Seeds of thale cress contained in siliqua
capable of launching them a significant distance.

Figure 20 – Poppy ovaries and seeds.

Figure 21 – Poppy seeds under grazing light.
Note their kidney-shaped appearance. L = 0.7 mm.

Figure 22 – Sow thistle (Sonchus oleraceus)
seed tufts. Diameter = 24 mm approximately.

 

A beautiful ovary to examine is that of the poppy. This has the shape of a covered jar (Figure 20) and is called a capsule. Above the ovary you can see the stigmas which are united to form a slightly rounded disc. When the ovary is mature, small windows open under the disc through which the seeds can exit. Collect a mature poppy ovary and beat it against the palm of your hand so that many of the small seeds come out (Figure 20). Examine the ovaries and section them to study their form and the arrangement of the seeds. Under the microscope, the poppy seeds show their alveolar kidney-shaped form (Figure 21). The wild pansy produces a fairly simple ovary which at a certain point opens in three parts letting the seeds fall. Oat seeds are equipped with two long appendages which contort when dried so that the seed falls to the earth penetrating it by rotating about themselves. The petunias instead produce small utricles that release miniscule seeds when overturned.

Nature produces seeds in such a variety of forms that you could even create a “herbarium” composed exclusively of seeds. If you prefer, this collection could be composed only of photographs and, why not, of drawings. Observe the plant seeds and describe differences in their aspect, colour and size.


METHODS OF SEED DISPERSAL

In many cases, the seeds fall at the base of the parent plant and the future generation finds little space for growth. To avoid this problem, many plants have developed different methods to disperse the seeds far away. One of the areas in which nature has gone wildest in the invention of diverse structures has been for the propagation of seeds. The dandelion, known also by the name of pissy bed due to its diuretic properties, transforms its yellow inflorescences into tufts, magnificent spheres composed of hundreds of miniscule parachutes (pappi), each of which transports a seed. If you look at a tuft under the stereoscopic microscope, you will note that the seeds are equipped with short upturned spines which serve to anchor the seed to the soil and facilitate penetration. The apple, the cherry and many other plants produce a sweet and colourful fruit which attracts animals to feed on it. Normally, the seeds are resistant to gastric juices and when the animal that ate the fruit defecates, the seeds are released into the environment. In the same way that nectar is a reward for the insects that pollinate the flowers, the fruit is a reward for the animal that carries the seeds away from the plant. Chestnuts and acorns are dispersed by squirrels and the acorns are often hidden in the ground where they are sometimes forgotten. The maple, lime and numerous conifers instead produce winged seeds that are carried away by the wind. Poplar seeds are covered with down which allows them to be easily borne a great distance by the wind.

 

Figure 23 – Flower of Geranium pratense.

Figure 24 – Ovary of Geranium pratense at diffe-
rent stages of maturation . Note how the pistil is
elongated and note the seeds at its base.

Figure 25 – Receding suddenly, the
laminae surrounding the style launch
5 seeds contained in as many sacs.

 

The plants which launch their seeds deserve a separate chapter. Their flowers transform themselves into true and proper launch mechanisms. This is the case of yellow sorrel (Oxalis corniculata), plants of the genus Geranium, thale cress (Arabidopsis thaliana) etc. There are many such plants and the devices that they have invented for carrying out this task are extremely diverse. Geranium plants (Figure 23) form five sacs at the base of the style each of which contains one seed (Figure 24). When mature, these sacs spring upwards, pulled by laminae that coil up (Figure 25), and the seeds are projected a distance of several metres. The thale cress has relatively long and cylindrical ovaries called siliqua (Figure 19). When these siliqua are mature and especially if a passing animal brushes against them, one of the two walls opens suddenly like a spring and shoots the seeds over half a metre away. The research and study of seed dispersal systems is particularly interesting and we will not be surprised to learn that collectors exist. Analyse different flowers, follow their transformation into fruit and study the seed dispersal methods. Do drawings and take photographs of their structure and of their transformations.


ROOTS

Delicately remove a plant from the earth. Brush away a little of the soil and wash off the rest so that the roots are well-cleaned. Cut off a small root and observe the root hairs under the stereoscopic microscope. Subsequently, you could make a transverse section of the tip of the root to observe at a high magnification. With the biological microscope, observe the transverse sections of the root (Figures 26, 27 and 28). On the exterior, you will encounter the root hairs, extensions of the epidermal cells whose nucleus is normally found in the hair. Attempt to identify the nucleus, if necessary using a nuclear dye to highlight it. The epidermis is the tissue that surrounds the root. At the centre of the section you can observe the central vascular cylinder or stele where the vascular vessels xylem (or wood), larger and more central, and phloem, narrower and peripheral, pass. The stele is surrounded by the endodermis which is composed of a single layer of cells. The endodermis regulates the passage of water and mineral salts absorbed by the roots. Just inside the endodermis is the pericycle tissue, a layer of cells capable of generating new roots. Between the endodermis and the epidermis there is tissue called the cortex, composed of parenchyma cells similar to that of leaves, but without chloroplasts. In some monocotyledons, instead, the vascular tissues form a cylinder around the central zone.

 

Figure 26 – Transverse section of a garlic root
(Allium tuberosum). Note the epidermis, the
cortex and the central vascular cylinder.

Figure 27 – Detail of the central vascular cylinder. Note the
endodermis, the pericycle tissue, the xylem and the phloem.

Figure 28 – Diagram of a root section.

 

The roots of a plant are often in symbiosis with the rhizome of fungi forming mycorrhiza. In many trees, the mycorrhiza are visible as a sheath on the outside of the root. In the herbacious plants instead the fungal hyphae penetrate the interior of the root. In this symbiosis, the fungi facilitate the absorbption of water and mineral salts by the plant, while the plant provides the fungus with sugars and organic substances.


STEMS

During the examination of the stems of herbacious plants and the young twigs of trees it is also useful to make transverse and longitudinal sections. Beginning at the outside, in a transverse section of the stem of a dicotyledon (Figures 29, 30 and 31) you will find in concentric layers: the cuticle, the epidermis, the cortex, the vascular cylinder (stele) and in some cases the pith (medulla). The cuticle is a thin layer impregnated with wax whose function is to reduce water loss. The epidermis can be composed of one or more layers of cells. In a green stem, the cortex is formed of live parenchyma cells. In the stele, the vascular tissues are found. The xylem vessels are orientated towards the inside of the stem and transport unrefined sap (water and mineral salts), whilst the phloem vessels are located towards the outside and transport processed sap (sugars and organic substances) from the leaves to the plant tissues.

 

Figure 29 – Transverse section of a sunflower
stem (Heliantus annuus), a dicotyledon.

 Figure 30 – Diagram of a section
of the young stem of a dicotyledon.

Figure 31 – Diagram of a stem section of a dicotyledon
prior to the commencement of secondary growth.

The stems of monocotyledons are not organised in concentric layers like those of the dicotyledons, but the vascular bundles are distributed throughout all of the fundamental tissue (“scattered” bundles, Figure 32). The pith generally contains reserve substances, but this can also be found desiccated and with the cells empty. Also in this case, in order to better identify the different parts of which the stem is composed and to understand their function, it is a good idea to read the corresponding charter of a biology textbook [001].

The longitudinal section of a stem is also useful for distinguishing the xylem vessels from those of phloem. In the vascular bundles, the xylem is located towards the inside and the phloem towards the outside. In the angiosperms, the xylem is composed of tracheids and tracheae (mature dead cells that can also form continuous vessels) and the phloem is formed of sieve-tube cells associated with companion cells. In the gymnosperms, the xylem is composed only of tracheids and the phloem only of sieve-tube cells.

Figure 32 – Transverse section of a corn stem.
In the monocotyledons, the vascular bundles are
dispersed throughout the tissue. (Field = 2.5 mm).

The support tissues collenchyma and sclerenchyma are also present in plant stems. The cells of collenchyma have thickened walls at the corners and are often located just inside the epidermis forming a continuous cylinder or distinct stripes. These cells support the young stems as they grow. Two types of sclerenchyma cells exist: fibres and sclereids. The fibres are elongated cells grouped into bundles which are often associated with vascular tissues. The sclereids are not elongated, are impregnated with lignin and sometimes with mineral salts which gives them their hardness and they often die when they reach maturity. These tissues are found in zones which have completed their primary growth.

 

Figure 33 – Transverse section of a Papyrus stem, Cyperus papyrus L.
(Cyperaceae). Dark background, ob 6 X. (Photo G.P. Sini).

Figure 34 – The same section under polarised light. The
sclerenchyma is bright in the photo. Ob. 6 X. (Photo G.P. Sini).

 

On the right in Figure 33, we see the external surface of a stem, protected by a compact layer of sclerenchyma bundles (hardened fibrous tissue for reinforcement). In the centre a vascular bundle with three large xylem tracheae is visible with the phloem immediately to the right. The vascular bundles are accompanied by reinforcing tissue (sclerenchyma, dark zones in a “C” shape that in the image (Figure 34) are bright due to their high lignin content which is birefractive). Polarised light can therefore be used to detect the sclereids.


MERISTEMS

A meristem is plant tissue consisting of undifferentiated cells in which new cells for mitosis are continually formed permitting the growth of the plant. The term meristem derives from the Greek merízein, which means “to divide" and recalls the process of cell division particularly active in these tissues. There are two principal meristems, that of the shoot (Figure 35) and that of the root (Figure 36).

 

Figure 35 – Apical meristem of a Hydrilla
verticillata
shoot in longitudinal section.

Figure 36 – Apical meristem of an onion root.

Figure 37 – Mitosis in an onion root
cell (anaphase). 400 X approx.

 

In the longitudinal section of the root you can see the apical meristem located near the tip of the root, in which the intense activity of cellular division which determines root growth takes place, and the root cap which protects the tip. In the root tip, it is possible to observe cells in various stages of cellular division or mitosis (Figure 37). For this purpose, take an onion and put it rooting in a glass of water. After a few days, when the roots are 4-5 millimetres long, cut off approximately 2 mm of the root tip and crush it. Put this piece in methylated spirits for about three hours, then place it on a slide and apply some nuclear dye such as 1% Toluidine blue for 1 - 2 minutes. If necessary, prolong the dying process. Wash away the excess dye and apply a coverslip. Now you are ready to search for cells undergoing mitosis. With a biology textbook, identify the stage of cell division of the various cells: prophase, metaphase, anaphase, telophase (look also at internet references [4005, 4006, 4007]).

This experiment can be best carried out with broadbean plants (Vicia faba) whose roots have large chromosomes. Allow some seeds of this plant to germinate in loose chippings or vermiculite (hydroponic culture).


LEAVES

The observation of transverse sections of leaf is very interesting (Figures 38 and 39). The leaves are the principal organs in which the process of chlorophyll photosynthesis takes place. Beginning from the upper surface of the leaf, we encounter the cuticle, the epidermis, the palisade mesophyll, the spongy mesophyll, the lower epidermis and the lower cuticle. The cuticle is a thin layer impregnated with wax. The epidermis is formed of one or more layers of cells. It is generally transparent and lacking in chloroplasts. The palisade mesophyll is formed from one or more layers of cylindrical cells arranged side by side. These cells are rich in chloroplasts which are clearly visible under the microscope and carry out an intense photosynthetic activity. The cells of the spongy mesophyll are irregular and arranged in such a way as to leave empty spaces useful for gas circulation. These cells are also rich in chloroplasts. Usually, chloroplasts adhere to the inner surface of the cell wall (Figure 40, the darker object is the nucleus). The lower epidermis is thinner than the upper epidermis. The surfaces of the leaf and in particular the lower surface are rich in stomata (Figure 41). The stomata are small openings, bordered by guard cells which regulate the amount of gas exchanged by the leaf and limit its water loss. In the centre of the leaf the vascular vessels pass and are called veins. On the undersurface of the leaf there are sometimes chambers or stomatal crypts containing hairs.

 

Figure 38 – Transverse section of a leaf.

Figure 39 - Diagram of a leaf section.

Figure 40 – Chloroplasts in the leaf cell of
a Bellis perennis. Diameter = 4 µm approx.
Objective Lomo apo 65X NA=1.1 imm. in water.

 

The stomata are best seen when observed from above (Figure 41) and not in section. Therefore, to observe the stomata, remove a piece of epidermis from the underside of a leaf. You will see not only the form of the stomata and of the guard cells, but you can also observe the form of the cells of that epidermis which often have the appearance of jigsaw pieces. In contrast to the epidermal cells, the stomatal cells contain chloroplasts. Observe in the photo also the nucleus (coloured red) which is also present in the guard cells.


EPIDERMAL AND HAIRY TISSUES

The examination of the epidermal tissues of the plant is also attractive since you can admire the different form of the cells. Many plants possess special hairs on the lower surface of the leaves and on the stems which are also interesting to observe. These hairs normally have the role of limiting water loss, but they also have other functions. To take samples of the epidermal tissue of a plant, use a pair of fine tweezers. Plants suitable for this type of observation are mullein, cinquefoil, Artemisia spp., Correa spp., oleaster, aubretia and all the plants with a velvety or hairy aspect. Some hairs have specialised points for secreting viscous substances or for injecting irritating substances into passersby (nettles). The sundew uses sticky hairs on the leaves to capture insects on which it then feeds. Some fruits are covered with hooked hairs which grasp to the fur of passing animals and so they are in this way disseminated far away (Figure 42).

 

Figure 41 – Stomata and epidermal cells of a leaf.

Figure 42 – Fruit of Cleavers Galium aparine
(Rubiaceae). Note the hooks, useful for
adhering to animal fleece. (Photo G.P. Sini).

Figure 43 – Leaflet of Salvia glutinosa
(Labiatae) with glandular digestive
hairs. (Photo G.P. Sini)

 

The petals of the small orange flower of anagallis are bordered with hairs which terminate in a small sphere. Hairs such as these are also visible on the leaves and stems of many plants, such as for example young rose or sage shoots (Figure 43). Search also for stinging hairs on nettles and describe them.

An epidermal tissue particularly easy to find in all seasons and which is very interesting to observe is that which covers the fleshy scales of the onion. Usually, this tissue is formed of a single layer of cells. This saves you from making a thin section of plant tissue, something which is rather difficult to realise. In this preparation, you can observe the shape of the cells, the primary cell wall, the nucleus and one or more nucleoli. If you are not used to doing this, it is very possible that, despite all your efforts, you will not be able to locate the nucleus of the cell. In fact, this is transparent and colourless and therefore barely visible. To make it more visible, a nuclear dye such as a 1% solution of Toluidine blue could come in useful. The nucleoli are where intense production of ribosomes, organelles destined for protein synthesis, takes place. The nucleoli appear as small discs inside the nucleus. Garlic also lends itself to this kind of analysis.

After having observed a plant tissue, it is interesting to compare the cells that compose a multicellular organism with the Protists, which are unicellular. You realise that while the Protists are free to move and to go wherever they wish, the cells of the tissues cannot do this. Not only this, but these cells are also very simplified with respect to those of the Protists and are transformed into specialised cells.


WET MOUNTS OF VEGETABLE TISSUES

 

Figure 44 – Equipment for carrying out wet mounts.

Figure 45 – How to obtain plant tissue
sections without using a microtome. 

 

Equipment to carry out wet mounts of vegetable tissues, Figure 44 from the left:

- 95 % alcohol in a hot water bain-marie (30-40°C);
- razor blade and elderberry pith;
- manual microtome, carrot and razor;
- box of microscope slides;
- box of coverslips;
- dropper;
- tweezers.

To examine plant tissues, you normally resort to sections. In this chapter, we will speak only about the realisation of wet mounts, that is, sections made by hand and if needed coloured and observed with the addition of water or alcohol, but not treated with the complex procedure necessary to realise permanent preparations.

Before beginning, procure the materials listed above. Some hours before starting, wash some slides.

To make the sections, take a new razor blade, expanded polystyrene foam or extruded polystyrene foam (not composed of beads). If you have difficulty in finding the polystyrene, use the elderberry pith. Cut the pith in half as though you were preparing a sandwich roll and place the tissue to be sectioned in the middle. With the razor blade, cut very thin sections, so thin as to be only one cell in width (Figure 45). As it is very difficult to achieve this width, you will have to make many attempts to practice the technique. Try also to obtain wedge-shaped sections so that at least in one area they have the correct thickness. This system lends itself to tender tissues such as leaves. Due to its greater hardness, the carrot is more suitable than the elderberry medulla for making sections of the small stems that are often rather hard to cut. To better hold the stems, make a "V-shaped” incision in the carrot.

Not indicated in the figure, but of great help in realising thinner sections, is a stereoscopic microscope with which you can better follow and if necessary correct the sectioning process. Observing the sectioning of the samples under the stereomicroscope and with a bit of experience you will be able to obtain sections even better than those possible with the manual microtome.

With a manual microtome such as that shown in Figure 44, it is possible to easily obtain sections, on condition that the blade is well sharpened. A manual microtome is not very expensive, but it is also possible to build one based on the principal of the differential screw for the precise progress of the sample, or you can use an outside micrometer caliper (1/100 mm) from which you have sawn off the fixed extremity.

Put the sections obtained on the slides, add some drops of water and try to eliminate any air bubbles present. Despite all your efforts, it is very likely that you will still have small air bubbles almost everywhere. To attempt to avoid this problem, allow some drops of methylated spirits to fall on the sections. Carry out this operation slowly to avoid contraction of the cells. Often, it is better to dye the tissues to highlight not only the nuclei but also to render the cells more visible. Prolonged immersion in alcohol tends to destroy the chloroplasts.


FOR THE LITTLEST ONES

As you know, children love playing. The experiments that follow are no longer microscopic observations, but games that you can propose to small children to tempt them to become interested in nature. If you approach children in the right way, it is possible to win their attention and curiosity.

The Cardinal
As you can see from the figures below (Figures 46 and 47), with a capsule and a bud from a poppy flower it is possible to create a cardinal.

 


 

Figure 46 - Poppy.

 

Figure 47 – How to make a cardinal out of some poppies.

 

Something well impressed on the mind
The poppy offers us another little game. This involves pressing a mature capsule on the forehead to leave an impression in the form of sunrays which remains for a few minutes.

The bladder campion goes pop!
Do you remember that flask-shaped flower that you collected in such a way as to prevent the air from escaping and which you then squashed on the back of your hand or on the forehead of a friend to hear "pop"? This is another Silene: Silene vulgaris, commonly known as the bladder campion. Play this game with a child and you will see how happy they are. This plant is also used as an ingredient in various dishes, for example "tagliatelle with bladder campion”.

Flower chains
Knotting some daisies together, you can make a daisy chain. You can also alternate different flowers. Twist some grass or make it into a plait, then fold it over and join the two ends. You will have a support into which you can insert many flowers to make a garland or a crown of flowers.


IN DEPTH

The subjects to study more in depth are:
The plants (Phanerogams) and their development (the plant cell, the tissues, the structure of the leaves, roots, stems, vascular tissues, meristems and the growth of the plant); the production of the principal organic compounds; the life cycle of the Gymnosperms and Angiosperms; reproduction of flowering plants (the flower, fertilisation, the ovule, the embryo, the seed, the fruit, seed dispersal); photosynthesis and respiration; the energy processes of the plant; the principal families of the Phanerogams and their characteristics. In the high school biology textbook indicated in [001] these subjects take up little more than a hundred pages.

As a guide to the observation of plant tissues and to recognise the structures that you will encounter, an anatomical plant atlas that contains photographs taken with the microscope will be very useful to you: [401] and [402]. Read also the accompanying explanations.

Botanical atlases based on drawings are also precious for the great quantity of explanatory diagrams they contain [003] and [004]. Of particular interest for the subject dealt with in this article are the tables which regard the plant cell, the structure of the stem and root, the various types of leaf, flower, fruit and seed of the superior plants.

For the identification of the different plants, you can refer to specific guides which contain photographs and/or drawings of the entire plant and its parts, such as flowers and fruit [404]. With guides such as these, you can manage to identify the genus and sometimes even the species of plant.

To continue further, you can refer to an university coursebook such as the following: [407], [410], or more recent.


CONCLUSION

The Phanerogams possess numerous specialised tissues and organs which are well adapted to terrestrial life. The variety of solutions adopted by different species to confront the problems of survival and competition with other species makes the study of these vegetable organisms particularly fascinating and complex. With the microscope, it is possible to carry out an infinite number of observations with the immense field composed of the Phanerogams. The indications offered here serve only as a first introduction to these plants. As you observe organs and tissues, you will find yourself faced with unknown structures, the identification and comprehension of which will require more in-depth knowledge.


BIBLIOGRAPHY

 

401 Kingsley R. Stern - James Bidlack - Shelley Jansky; Introductory Plant Biology; McGraw Hill Higher Education; pag 593; 2008.
402 Gerlach D., Lieder J.; Taschenatlas zur Pflanzenanatomie; The microscopic structure of vascular plants. An atlas rich of drawings and pictures; pag 150.
403 AAVV; Nature lower's library field guide to trees and shrubs;  The Reader's Digest; London 1981; pag 304
Atlas to identify the herbaceous plants, provided with a lot of drawings and pictures.
404 AAVV; Nature lower's library field guide to wild flowers; The Reader's Digest; London 1981; pag. 448.
atlas to identify the woody plants, complete with many drawings and pictures.
405 Strasburger E.; Strasburger's Text-Book of Botany; 895 pag., Longman Group United Kingdom;
A college textbook complete with many drawings.
406 Ruzin, S.; Plant Microtechnique and Microscopy; Oxford University Press Inc, USA 1999, 57 figs, 334 pag.
A superb modern reference book, full of practical information, well written and designed, but of limited use to the amateur microscopist.
407  
408  
409  
410  
411 White J.; Pollen, its Collection and Preparation for the Microscope; Northern Biological Supplies Ltd.; pag. 40

Look also at the works and Internet references of general character indicated in the presentation article of this guide.


INTERNET RESOURCES

4001 -
4002 -  
4003 -
4004 -
4005 - http://www.biology.arizona.edu/cell_bio/activities/cell_cycle/cell_cycle.html  Onion root tips
4006 -
4007 - http://www-saps.plantsci.cam.ac.uk/worksheets/ssheets/ssheet17.htm  Mitosis in root tips
4008 - http://www.floralimages.co.uk/  Floral Images
4009 - http://florenligne.free.fr/index.htm  Flore en ligne
4010 -
4011 - http://www-saps.plantsci.cam.ac.uk/index.htm  Sience and Plants for Schools
4012 - http://www.publicbookshelf.com/public_html/Methods_in_Plant_Histology/  Methods in Plant Histology
4013 - http://www.ou.edu/cas/botany-micro/bot-linx/  Botanical Links
4014 - http://www.microscopy-uk.org.uk/mag/indexmag.html?http://www.microscopy-uk.org.uk/mag/artmar04/jbbotany2.html Botanical Microtechnique Part 1
4015 - http://www.microscopy-uk.org.uk/mag/artmar04/jbbotany2.html Botanical Microtechnique Part 2
4016 - http://www.pollenwarndienst.at/index.php? Pollen Atlas
4017 -

Internet keywords: college textbook botany, reference book botany, atlas botany, botanical microtechnique, botanical histology.

 


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