Procedure for Exercise B

  1. Obtain a leaf of aloe, the burn plan. Note that the leaves are fleshy, or succulent. This is commonly seen in plants in arid environments as an adaptation for water storage. Break open a leaf. Note the gel in die middle of the leaf. This contains aloin and chryso-phanic acid, ingredients that have been known for centuries to heal the skin. Many people keep an aloe plant in the kitchen window and apply the gel to minor burns and cuts. Aloe is the main ingredient in many over-the-counter first-aid crcams and in Oil of Olav, a famous cosmetic skin conditioner.
  2. Fold the leaf so that the translucent epidermis (it resembles clear plastic wrap) extends over the torn edge. With a forceps, remove the translucent epidermis to reveal a layer of green with splotches of white. This layer is above the gel of the leaf. Using a pair of forccps, take a small piece of this green layer and place it on a slide. Add a drop of distilled water, and cover with a covcrslip.
  3. Scan under scanning (4 x objecdvc) or low (10 x objective) power for a section of only one or two layers thick. Make sure you focus on the top layer, not the in-between layers. Focus on a small group of cells. Bring to high power (40 x objective). Note that the plant cells have a regular shape. They look like rectangles (fig. 1.1). The outline of the cell Ls clear because

Chloroplast s

Cell Wall

Nucleus

Chloroplast s

Cell Wall

Nucleus

Elaioplast Avocado

FIGURE I.I PLANT CELLS FROM THE MESO-PHYLL OF AN*ALOE LEAF.

the outermost boundary of a plant cell is die cell wall. The cell wall is rigid and gives support to die cell. In these cells, the primary wall is composed mainly of die polysaccharide cellulose. In some types of plant cells, additional layers arc laid down inside the primary wall. This is the secondary wall, which is much thicker and contains lignin in addition to the cellulose. The strength and character of wood is due to secondary walls. Secondary walls and the cells in which they are found are discusscd in both Laboratory Topic 3 and Laboratory Topic 15.

The cohesiveness of plant tissue is due to the middle lamella, an intercellular glue composed of pectins, that holds die cell walls of neighboring cells together. Pcctinase was used in Exercise A to free individual cells from the middle lamella. The middle lamella is most pronounced when die corners of plant cells adjoin.

  1. View the interior of one of these cells. Note the spherical bright green discs that appear to hug the interior side of die cell wall. These arc chloroplasts, and die green pigment contained within them is chlorophyll. Chloroplasts arc die sites of photosynthesis, in which the energy of die sun drives the joining of water and carbon dioxide to make glucose. The process of photosynthesis is further discusscd in Laboratory Topic 5. i Although not visible, a plasma membrane encases the protoplast. The plasma membrane is a lipid bilayer with embedded proteins. It is selectively permeable and regulates what passes into and out of die cell. You could dearly sec the plasma membrane in Exercise A.
  2. Scan several cells to spot a nucleus. The nucleus is a perfect sphere and clearly outlined by the nuclear envelope, a double membrane layer. It is larger than a chloroplast, and it may be adjacent to the wall or somewhere in the center of the cell. The interior is granular because of chromatin, the form DNA (deoxyribonucleic acid) takes in a nondividing cell. DNA is the genetic blueprint of life.

You may also see one or two smaller circular regions within the nuclcus. ITiese arc nucleoli (sing., nucleolus). The nucleolus is rich in RNA (ribonucleic acid), and is involved with protein synthesis in die cell.

  1. Although the interior of the cell looks empty, that is really not the case. The center is occupied by the large ccntral vacuole. The tonoplast is the vacuolar membrane and will not be visible. The vacuole serves as a reservoir for materials, including water, that the cell can draw upon when needed. It may also be a place to sequester waste products. In iact, some products accumulate in such great quantities that they precipitate out as crystals. You may have seen single or bundles of ^needles" as you scanned your slide ofaloe. These are crystals and arc discussed further in Exercise D. Some plant pigments are also stored in the vac uole, giving color to truits and flowers. These pigments have been used for millennia to dye cloth, much as you will do in Excrcisc E.
  2. Plant color is also imparted by another organelle, the chromoplast. Chromoplasts contain the carotcnoid pigments of yellow, orange, and red. Chromoplasts are related to chloroplasts; in fact, both originate from a common precursor called a proplastid that gives rise to all of the plastids. Plastids include not only chloroplasts and chromoplasts. but Icucoplasts. Leucoplasrs arc colorless and include amyloplasts, which accumulate starch grains, and elaioplasts, which accumulate lipids. Amyloplasts arc ilirthcr discussed in Laboratory Topic 13.

To see elaioplasts, obtain a small piece of avocado fruit and smear to a thin layer on a slide. Add a drop of distilled water and a drop of Sudan III solution. Sudan III will stain the lipid containing elaioplasts. Focus first under scanning (4 x objective) or low 110 x objective) power and then bring on up to high power (40 x objective). What color do the elaioplasts stain in die presence of Sudan III? Draw a tew elaioplasts in the space following:

Chromoplasts conic in a variety of shapes and sizes. You will know them by their color and abundance in certain fruits, flowers, and in some cases, roots. To view a chromoplast, rake a small piece of the skin of a yellow; orange, or red bell pepper. Place it on a slide, add a drop of distilled water, and cover with a coverslip. Focus first under scanning (4 x objective) or low 10 x objective) power and then bring up to high power ¡40 x objective). Draw some representative chromoplasts in die following space:

Look closely at the cell wall of the pepper skin. Can you spot the cytoplasmic bridges through the walls? These are the plasmodesmata (fig. 1.3) that interconnect plant cells.

Prepare slides as before, and look for chromoplasts in the skin of a tomato, the root of a carrot, or the petals of a marigold. Draw some representative samples in the following space:

Plasmodesmata Nucleus

Vacuoie

Plasma membrane Middle lameiia

Chromoplasts Tomato Labeled Drawing

Grana CnloropJasi

Mitochondrion

Ceil wall

Pollen Mikro

FIGURE 1.2 SCANNING ELECTRON MICRO-GRAPH OF POLLEN GRAINS OF AMARYLLIS.

FIGURE 1.3 TRANSMISSION ELECTRON MICROGRAPH OF A PLANT CELL.

EXERCISE C: Plant Ultrastructure

Many of the details of the contents of a plant cell only became discernible with the development of the transmission electron microscope (TEM > in the 1950s. In microscopy, two properties must be considered: magnification and resolution. Magnification tells how much larger the image is compared to the actual size of the specimen. Resolution is the ability to disccrn fine details— in other words, to distinguish two closely spaced objects as two separate points. Resolution is inversely proportional to the wavelength of radiation used in a microscope Electron microscopes replace the light beam of the compound microscope with an electron beam. Since the wavelength of an electron is much smaller than that of visible light, the electron microscope has at least a thousandfold greater resolution than the light microscope.

The scanning electron microscope (SEM) scans an electron beam across the surface of specimens that have been coated with a thin layer of gold. This produces a three-dimensional image of plant structures. In this exercise, you will view both scanning and transmission electi on micrographs and learn the ultrastructurc of a plant cell.

Materials Needed for Exercise C

Scanning electron Transmission micrographs of electron micrographs plant structures of plant cells

Plasmodesmata Nucleus

Vacuoie

Plasma membrane Middle lameiia

Procedure

  1. Examine figure 1.2 as an example of the three-dimensional image possible with a scanning electron microscope. View other scanning electron micrographs that arc available in the laboratory.
  2. Using table 1.1 and figure 1.3, identify and label plant cell ultrastructure in worksheet 1-2, figure 1.5 at the end of this laboratory topic.

FIGURE 1.2 SCANNING ELECTRON MICRO-GRAPH OF POLLEN GRAINS OF AMARYLLIS.

Grana CnloropJasi

Mitochondrion

Ceil wall

TABLE I.I PLANT CELL ULTRASTRUCTURE

J Cellular Structure

Description Checklist

Cell wall

Rigid outer structure that supports and protects primary wall of cellulose; in some plants, a secondary v/all reinforced with lignin forms

Chloroplasts

Double membrane-bounded organelles: site of photosynthesis; chlorophyll (green) plastids. internal organization of grana. membranous stacks, and stroma, protein-rich matrix

Chromoplasts

Pigmented plastids of yellows, oranges, and reds: impart color to fruits, leaves, and flowers

Cytoskeleton

Scaffolding of microtubules and microfilaments; gives shape and support

Cytosol

Matrix of the cell, bounded by the nucleus and plasma membrane

Endoplasmic reticulum

Network of membranous canals through the cytoplasm; functions in transport; lipid synthesis associated with smooth ER; rough ER studded on its free surface with ribosomes. associated with protein synthesis

Golgi apparatus

Stack of flattened membranous stacks; site of packaging, modification, and secretion

Leucoplasts

Colorless plastids; include starch-storing amyloplasts and lipid-storing elaioplasts

Microbodies

Membrane-bounded organelles; location of certain metabolic reactions

Middle lamella

Intercellular material that glues adjacent plant cells together; composed of pectins

Mitochondhon

Double membrane-bounded organelle: site of aerobic respiration: cristae are folds of the inner membrane, matrix is the compartment enclosed by the inner membrane

Nucleolus

One or more dark-staining areas in the nucleus: RNA-rich regions involved in protein synthesis

Nucleus

Control center that contains genetic blueprint of DNA in the form of granular chromatin; surrounded by nuclear envelope, a double membrane with small openings, the nuclear pores

Plasma membrane

Lipid bilaycr with embedded proteins; governs the movement of materials into and out of the cell

Pfasmodesmata

Cytoplasmic bridges that cross cell walls and connect neighboring plant cells

Ribosomes

RNA-rlch organelles; site of protein synthesis; may be found free in the cytoplasm or attached to ER

Vacuole Very large, centrally located organelle bound by a tonoplast membrane;

depository for storage, wastes and pigments (reds and blues)

EXERCISE D: Crystal Persuasion

Plants do not have an cxcrctorv system, and therefore some

substanccs have the potential to accumulate lo toxic levels. By forming crystals that precipitate out in the cell vacuoles, such substances arc taken out of solution, thereby becoming inert and harmless. In tact, stored crystals are often quite beautiful. They have also been used for identification purposes in forensic science and archaeology. Most contain calcium, cirhcr calcium oxalate or less commonly; calcium carbonatc. Phvtoliths arc crystals found in »' • -

the walls of ccrtain families and genera of plants, usually monocots. They are made of silica (monosilicic acid) that is dissolved in the water absorbed by the roots. Plants cannot use silica, so often it is deposited in cell walls. If the plant dies or is cooked, the silica deposits survive intact for millenia, and because specific groups of plants produce uniquely shaped phytolirhs, they have been used in archaeology as an identification tool. In this exercise, you will discover the types of crystals found in several plants.

6 Appijfd Botam

Cystolith Lithocyst

FIGURE 1.4 PLANT CRYSTALS. (A) RAPHIDES IN PINEAPPLE. (8) DRUSE IN LEAF CELLS OF ELEPHANT EARS. (C) CYSTOLITH IN LEAF CELLS OF RUBBER TREE (FICUS ELASTIC A).

FIGURE 1.4 PLANT CRYSTALS. (A) RAPHIDES IN PINEAPPLE. (8) DRUSE IN LEAF CELLS OF ELEPHANT EARS. (C) CYSTOLITH IN LEAF CELLS OF RUBBER TREE (FICUS ELASTIC A).

Materials Needed for Exercise

Glass slides

Compound light microscope

Covcrslips

Dissecting needles

Dropper bottles of distilled water

Elephant's ear

(Colocasia ultissima)

Pineapple, can ofcrushed

Razor blades, single-edged

Rubber tree (Fiats clastic a)

Procedure for Exercise D

  1. Take a small sample of crushed pineapple and spread it thinly on a slide with a drop of distilled water. Cover with a covcrslip. View first under scanning (4 x objec tivc) or low (10 x objective) power. Look for the bunches ofnccdlclike crystals (tig. 1 Aa ». Focus under high power (40 x objective) for a closer look. These are called raphides. Composed of calcium oxalate, they are common in many plants. Recall that you saw these crystals in the leaf cells of aloe viewed in Exercise B. You may also find these crystals in the leaves of dumbcane (DieffcnbaeJria). This common houscplant got its nickname because, if the leaves arc eaten, the sharp raphide crystals stab the tongue and throat, causing inflammation to such an extent that the person is struck dumb or mute. Ifdieffenbachia is available, tear up a piece of leaf tissue and prepare a slide for viewing under the microscope. Sketch some representative raphides in the following space:
  2. Another crystal of calcium oxalate takes the form of a star or druse tig. I Ab). To view a druse, tear up a piccc of elephant's car. Place it on a slide with a drop of distilled water, cover with a covcrslip, and view first under scanning 4 x objective) or low (10 x objective power to scan for the druses. Bring up to high power (40 x objective) for a closer look. Elephant's car is taro, the source of poi. the native dish of Hawaii. Poi Is prepared from the underground stem of taro, but the leaves are also eaten as foods are wrapped and cooked in taro leaves. In fact, the word luauy which has come to mean a Hawaiian style feast, is the native word for these leaves. Sketch a druse in the following space:
  3. A cystolith (cell rock) is another type of plant crystal. It is composed of calcium carbonate and is present in a small number of plant families, including that of the rubl>er tree. Since these leaves are quite stitY, you should be able to cut a small piece of thin cross section of a Ficus clastic a leaf without too much trouble. Place your thin section on a slide, add a drop of distilled water, and cover with a covcrslip. Scan under scanning (4 x objective) or low (10 x objective) power to search for the colorless epidermal cclls where the cystoliths arc located in lithocysts (rock cells). Focus under high power (40 x objective) when you spot what looks like a hanging bunch of grapes (fig. 1.4c). That's the cystolith! Skctch a cystolith in a lithocvst in the following space:

EXERCISE E: Plants to Dye For

Flowering plants brought color to the green world. In mosses, ferns, and gymnosperms (e.g., conifers), the predominant color is green, but in the angiospcrms, a variety of other colors have been added to the palette. Vibrant or subtle, these colors shimmer from fruits, petals, seeds, and even some leaves. It is no wonder that humanity long ago created methods to extract and enhance these colors from the plant rainbow.

For most of human history, dyes for cloth and cosmetics were obtained from natural sources, either vegetable, animal, or mineral. Most of these natural sources were replaced by die synthetic aniline dyes first obtained from coal tar dyes during die latter half of the nineteenth century. Lately, i hough, there has Ixren renewed interest in vegetables dyes, which are admired for their subtle and unique hues. In this exercise, you will learn about the chemistry of plant dyes and extract dyes from a variety of plants.

Locate another cell structure prominent in these cells. It is spherical and centrally located. Note that it has one or two small, dark-stained circles within it. What is it?

Materials Needed

Beakers, 50 ml

Beakers, 250 ml

Compound light microscope

Coverslips

Dropper bottles of ammonia

Dropper bottles of distilled water for Exercise E

Dropper bottles of vinegar

Forceps Glass slides Hot plate Marker pens Natural wool yarn, mordanted with alum

Red cabbage leaves

Procedure for Exercise E

1. Peel off the epidermis of a red cabbage leaf by folding a small section of a leaf in half and grabbing the thin purple layer with a forceps. Place a small piece in a drop of distilled water on a glass slide and cover with a cov-erslip. Examine first at scanning (4 x objective) or low

10 x objective) power and then bring up to high power (40 x objective).

  1. Note that the interior of each cell is purple. The purple color is due to the presence of a pigment called anthocyanin, which is contained within the vacuole of the plant cell. Note also the ccll w alls that define the boundarv of each cell. Scan around. Do vou see the prominent crystals in the vacuoles of many of the cells? Using figure 1.4 as a guide, identify the type of crystal vou sec.
  2. Anthocyanins have been used for millennia as a source of natural textile dyes. They also have some unique chemical properties that you will be investigating.
  3. Tear up 1 red cabbagc leaf, and place it in a 250-ml beaker, add just enough distilled water to cover. Also add to die beaker a length of undyed wool that has been treated with die mordant alum and simmer for 30 minutes on a hot plate. (.-Mum, or potassium aluminum sulfate, is a mordant, a chemical agent that makes the dye adhere to the fabric. It can be found in the spice section of a supermarket.) The color of die dye may change, depending on whether a mordant is used or not, and if the fabric is treated with different mordants.
  4. L'se the same procedure to set up other dye vats and wool. Some plants to try include strawberries, blackberries, blueberries, cranberries, beets (the pigment here is betacyanin), yellow onion skins, red onion skins, mullein leaves, Indian blanket flowers (Gail-lardia pitlcbc/la)y or any other plant of your choosing. Record your findings in worksheet 1-2 at the end of this laboratory
  1. After you have completed dyeing die wool in the red cabbage juice, decant the anthocyanin solution. Pour off equal volumes of this solution into tw o 50-ml beakers. Label one beaker "A" and the other "B."
  2. Add a few drops of ammonia to beaker A. What color change do you see? Add a few drops of vinegar to beaker B. What color change do you see?

Andiocyanins are natural pH indicators in that, in the presence of acid or base, they undergo a chemical change I hat is visible in color. The pH scale is used to measure the acidity or alkalinity of substances. It ranges iTom 0 to 14, with 7 being neutral, the lower end of the scale is add, and the higher end is alkaline or basic. In the presence of acid, anthocyanins give up hydroxy ions (OH ), resulting in a red color. In the presence of a base like ammonia or baking soda, they pick up hydroxy ions, resulting in a blue or green color. How could you reverse the color changes?

TERMS TO KNOW

acidity 7

mitochondrion 5

alkalinity 7

mordant 7

amyloplast 3

nucleus 3

anthocyanin 7

nucleolus 3

base 7

nuclear envelope 3

ccllulase 1

pecrinase 1

cellulose 2

pH scale 7

cell wall 3

phytoliths 5

chlorophyll 3

plasma membrane 3

chloroplast 3

plasmodesmata 3

chromatin 3

plastids 5

chromoplast 3

primary cell wall 2

cvstolith 6 •

proplasrid 3

cytoplasm 5

protoplast 1

DNA

raphides 6

(deoxyribonucleic

resolution 4

acid) 3

RNA (ribonucleic

druse 6

acid) 3

elaioplast 3

scanning electron

hydroxy ions(OH * ) 7

microscope (SEM) 4

leucoplast 3

sccondarv cell wall 2

lignin 2

succulent 2

lithoevsts 6 •

tonoplast 3

magnification 4

transmission electron

middle lamella 3

microscope (TEM) 4

microbodies 5

vacuole 3

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Responses

  • bilba
    What is the cytoplasmic bridges on a pepper skin called?
    8 years ago
  • Roxanna
    Why add water to aloe leaf on microscope slide?
    8 years ago
  • Christina
    What color do avacado elaioplasts stain with sudan III?
    8 years ago
  • JOHN
    What color do the elaioplasts stain in the presence of Sudan III?
    7 years ago

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