PROJECT: INVESTIGATION OF PLANT STRUCTURE

For introductory biology for science majors

Department of Biology, Southern Oregon State College, Ashland, OR 97520

OBJECTIVES

• Describe plant structure and its relation to function, particularly energy use, and adaptations to habitat.
• Apply information and techniques learned in class to analyze a plant that you select.
• Prepare a formal laboratory report in format of a scientific paper

Sample Calendar

• Oct 19 Introduction to project. Discuss questions about exercise. Select plant.
• Oct 24-Nov 3 Do laboratory work. Make observations, hand section and stain leaves and stems, prepare diagrams, drawings. Write paper; find references
• .Nov. 4 Saturday 10-4 Open labs for plant project work.
• Nov. 9 Semi-final draft due at beginning of your lab Peer evaluations IN LAB Drafts and comments will be returned to you that day in lab
• Nov. 16 PLANT REPORTS DUE

PROCEDURES

Any plant with leaves and stems available over a two to three week period will be suitable. You may use a house plant available from a local store, the SOSC greenhouse, or an outdoor plant (not deciduous), native or cultivated. Plants should be approved by instructor. The plant should be identified with its scientific and common names. Plants used as examples in your text cannot be used.

B. Techniques

Techniques used in the study of leaf and stem anatomy include those described in Microscope Methods section. After materials have been sectioned and stained, drawings and diagrams should be made as soon as possible. You will probably need to make several slides of the same structure before you get a good preparation that will reveal your plant’s innermost secrets.

Temporary slide mounts can be made semi-permanent by drawing off excess liquid with a paper towel and then sealing the edges with clear fingernail polish. These preparations will last for several weeks if edges are completely sealed. Keep slides flat.

Another method for making temporary mounts semi-permanent is to carefully remove the cover slip, draw off as much of the liquid as possible and then add a drop of glycerine. Carefully lower a new cover slip over the specimen. Remove excess glycerine from the edges of the cover slip and seal them with clear fingernail polish.

C. Descriptions of plant parts

Discover as much as you can about your plant. The following questions may serve as a guide in your studies of morphological and anatomical features of your plant.

1. Gas exchange

• Where are stomata located?
• On which surfaces?
• What are the shapes of cells in the epidermis (guard cells, subsidiary cells, ordinary epidermal cells)?
• How many stomata are there per square millimeter?
• Do you know the diameters of the fields of view?
• Are stomata located in depressions?
• What is the nature of the cuticle (thick or thin)?
• Are epidermal hairs present?
• What is their structure?
• Is the epidermal layer unicellular or multicellular?

2.Transport system

• What is the distribution pattern of vascular bundles?
• What is the arrangement of xylem and phloem in each bundle?
• What types of cells are present in these two tissues?
• Has secondary growth occurred in the stem?
• How are veins arranged in the leaf?
• What is the pattern of venation?

3. Leaf and photosynthesis

• Where is chlorophyll located in the leaf?
• How much chlorophyll is present on a mg/g basis?
• Which photosynthetic pigments are present?
• Is there chlorophyll in the stem?
• What types of cells make up the mesophyll? cortex? pith?
• Does mesophyll in the leaf occur as both palisade and spongy?
• How many layers of palisade parenchyma are there?
• Are crystals present? Where?
• Is there collenchyma or sclerenchyma in any ground tissues?
• Is starch present in any cells?
• Are there non-green parts? Where is starch stored?
• If starchy parts are kept in dark, does starch disappear?
• Is the leaf anatomy that of a C3, C4 or CAM plant? The acidity of CAM plant cytoplasm changes during the day. If your plant appears to have CAM anatomy, test the acidity of leaf juices after a dark period and after 24 hours of light.

4. Special adaptations

• Does your plant have special features or adaptations that relate to functions or the habitat (xerophyte, hydrophyte, mesophyte) where the species is native?
• Is it protected from desiccation or from predation?

REPORT

THE FORMAT DESCRIBED IN THE APPENDIX, HOW TO WRITE A LAB REPORT IN THE FORMAT OF A SCIENTIFIC PAPER, MUST BE USED.

In the Introduction, include general information about the plant, common and scientific names, native habitat, uses (medicinal, herbal, etc.). Cite sources of information using the format described under REFERENCES. Describe the appearance of the plant and the arrangement of parts as seen by eye (tall, shrubby, opposite pinnately compound leaves, etc.). Point out any interesting features or information about the plant. Pose any special questions about the plant that you are going to attempt to answer in this study. Results section will include observations, descriptions, and illustrations including a habit sketch that shows arrangement of leaves on the stem and the general growth pattern of the plant (do not include the pot in your sketch), cross-sections of leaf (including vein and mesophyll) and stem, and epidermal view of leaf and possibly of stem. Illustrations can involve both diagrams and drawings, but must include drawings. Additional drawings of special features (stomata, epidermal hairs, etc.) should also be included. Include magnifications for each illustration along with titles and labels.

Results must include written descriptions of the illustrations. Based on macroscopic and microscopic observations, describe in some organized way what you observed. Refer to diagrams and drawings. All descriptions must be given in words. The picture does not speak for itself. You must describe it.Number figures in the order of their description in the results. Group figures on pages separate from the prose. Each figure should have a number and title e.g., Fig. 1. Transverse Section of Stem Showing Vascular Bundle, Phloroglucinal Stain, and be completely labeled (regions, tissues, cell types, specific structures). Figures may be cut out and pasted in groups on a clean sheet of paper when you are putting your final report together. For the discussion, summarize or synthesize your most significant findings. Relate your findings to what you know or can deduce about the plant's habitat. If you obtained information from the literature, cite your sources. If you asked any particular questions in the Introduction, answer them in this section.

Find at least two botanical references (in addition to your textbook). These may be books on plant physiology, plant anatomy or morphology, leaf or stem structure or physiological ecology. In your paper, where you discuss ideas or observations from other sources, cite them using the format described in the Appendix, How to write a lab report.SLIDES: When you turn in your report, include a representative prepared microscope slide for each illustration in your paper. Place a label on each slide indicating your name, plant, type of section, stain.Turn in peer reviews with final copy of paper.

MICROSCOPE METHODS

I. PREPARATION OF SLIDES

A. Temporary or wet mounts

2. Place a small portion of material in the drop of water. If material is filamentous, tease it apart with a pair of dissecting needles.

3. To avoid trapping air bubbles when adding the cover slip, place the edge of the cover slip on one side of the liquid and then lower it gently onto the drop. This forces air bubbles away from the material.

4. If water accumulates around the edges of the cover slip, soak it up with a piece of paper toweling touched to the edge of the cover slip. If additional water is needed, add a small drop to the edge of the cover slip. If the slide starts to dry while being viewed through the microscope, water may be added in a similar fashion.

B. Hand sectioning

Slide method

2. Hold the object in place on the slide with the tip of the forefinger of the left hand.

3. Hold a single-edged razor blade firmly with the right hand. Steady the razor blade by resting its flat surface against the tip of the left forefinger.

4. Carefully slice through the section.

5. Repeat the procedure without moving the left forefinger but simply applying more pressure with the razor blade against the tip of the left forefinger. As each section is cut, it should float onto the drop of water before you make the next slice. As continued sections are cut, each will be thinner than the preceding due to increasing pressure being exerted against the tip of the left forefinger with the razor blade.

Hand-held method

2. Hold the single-edged razor blade in the right hand and place the flat side of the razor blade against your index finger and the edge of the razor blade against the sample material.

3. Cut sample with a slicing movement of the blade as you pull the blade across the sample towards you. Try to keep the slice as thin as possible. Angling the section in an upward direction will yield a section with one edge thicker than the other.

4. Place the section in a drop of water on a clean slide. 5. Rotate material 180 degrees and prepare another thin section from this opposite side.

6. Repeat this technique until you have several thin sections.(If you are left handed, hold the specimen with your right and cut with your left.)

Tips for successful sectioning

• Use a sharp razor blade.
• Make many sections. It is rare to obtain a perfect section on the first slice. It saves time to make several sections before adding a cover slip to the preparation.
• Make slices that taper in thickness. If the section tapers, it may be too thick near one end and perhaps too thin near the other end, but somewhere in between it should be usable; that is, thick enough to preserve the structure but thin enough to transmit light.
• Do not attempt to make a complete section of a large object.
• Thin, flat objects such as leaves should be rolled to about pencil thickness and then sectioned across the end of the roll.

C. Epidermal peels and epidermal impressions

1. To prepare an epidermal peel, hold the margin of a leaf firmly with the left hand between thumb and forefinger.

2. With the right hand, begin to tear the leaf in half by pulling the right side toward you. As you tear the tissue, begin to pull the right side to the right (askew) of the original line of the tear.Along the margin of the tear, there will be a thin layer of the epidermis usually only one cell thick.

3. Place the epidermal peel in a drop of water on a slide.

4. Cut off any untorn section of the leaf and discard. Place a cover slip on the material and examine.

An epidermal impression can be obtained from thin or fragile leaves whose surfaces do not yield good epidermal peels.

1. Wipe the leaf surface with 50% ethanol, then apply several layers of clear fingernail polish to the surface, allowing each application to dry before the next is applied.

2. After two or three applications, peel off fingernail polish, mount it in water on a clean slide, and cover with a cover slip. Epidermal cells leave an impression in the polish.

D. STAINING

TOLUDINE BLUE is useful for identification of cell types and tissues. Sections are collected in water, immersed in an 0.05% aqueous solution for 10-60 sec, rinsed in water for 1-2 min., and mounted in water. Lignified walls and structures containing tanins stain blue-green or light green. Walls of collenchyma, parenchyma, sieve tube members, and companion cells stain pinkish-purple.

IODINE (IKI) is used to identify starch in cells and can be used to differentiate other structures. Starch stains blue-black. Immerse sections in stain for 2-4 min., then transfer to water or 50% glycerine.

The purpose of an illustration is to record observations and to identify and understand structures and their functions. Illustrations summarize information that otherwise needs many words to describe. In preparing illustrations, emphasize accuracy, neatness and scale. Two types of illustrations are diagrams and drawings.

A. Diagrams

B. Drawings

C. Guidelines for Illustrations

• Make illustrations only after careful study of the subject. Plan drawings and diagrams before you start.
• Be neat. Erase neatly and completely.
• Illustrations should be LARGE. Show structures in proper proportions with no crowding of parts and labels.
• All illustrations must have a title which identifies the name of the plant, the part being drawn (stem, leaf, root) and type of section (cross, radial, tangential). The magnification of the illustration accompanies the title.
• Position the drawing slightly to left-of-center of the paper with labels on the right.
• Label the illustration completely. Begin pointer lines ON the designated object and make these lines parallel when possible. Never cross pointer lines. Label each pointer line fully.
• Use biology drawing paper. Draw on one side of the paper.
• Use a sharp, 3H (or equivalent) drawing pencil. Make clean, sharp lines. Plain line illustrations tell the story without the distraction of artistry.

  D. Magnification of an illustration

  Magnification of a diagram or drawing of an object in the microscope is equal to the magnification obtained with the microscope ONLY if the drawing is the same size as the object appears in the microscope. This rarely happens. Magnification of the object in an illustration which has been enlarged over that seen by the microscope can be calculated by the following:

  Magnification = SIZE OF DRAWING/ACTUAL SIZE OF OBJECT

  The size of the drawing of the object is obtained directly by measuring its size on the paper. To determine the actual size of an object as seen through the microscope, refer to the explanation above. Field diameters are approximately 4.0 mm (4X), 1.5 mm (10X), and 0.35 mm (43X).

  APPENDIX

  HOW TO WRITE A LAB REPORT IN THE FORMAT OF A SCIENTIFIC PAPER

  A scientific paper is a method of communication, a way of telling others about data that you have collected and what you think the data mean. The paper requires correct use of language. The report should be written in a clear, concise manner so the reader can easily understand what you have done. There should be enough detail so that another person could repeat your work.

  Rules for writing a scientific paper are rigid (Gubanich 1977). It takes time and practice to become familiar with the style required. These instructions apply to papers in many scientific journals.

  The TITLE should enable the reader to tell what you have done without having to read the paper. A self-explanatory title usually tells the factor that was manipulated, the major parameter that was measured and the organism that was used.

  SUMMARY.

  In a few sentences state (1) the problem being investigated, (2) the approach or methods, (3) the results and (4) the conclusion. It is only one paragraph.

  INTRODUCTION.

  State the problem and the background out of which a question arises. What is the purpose of this paper? What observations or ideas led to this particular experiment; what research has been done in the past? Describe the significance of the problem. If we knew the answer, what would it tell us? Develop a hypothesis or tentative explanation that guesses the answer. Include your rationale for this particular hypothesis. Background information from other sources (textbooks, journals, etc.) must be cited and listed in the Literature Cited section. The introduction should have no figures.

  METHODS AND MATERIALS.

  Explain in detail how you did the work. Describe the experimental design, the apparatus, methods of gathering data and controls used. Include enough detail about materials and procedures so that another student could duplicate your work. Do not include the rationale for the work nor any interpretations. Do not use a chronological approach (First, I tried this, then that...), but organize the methods in a logical order.

  Give names of materials, their sources, how you handled the material, relevant temperatures, concentrations, and times. Explain how the work was done. If you cut a plant or animal or applied a substance, tell exactly how and where on the organism the work was done.

  The Methods and Materials section and subsequent sections of the paper should be written in past tense. A scientific paper is a report on something that you did in the past. Do not make a list of equipment like a shopping list and do not give instructions as in a lab manual or a recipe. Report what you did.

  RESULTS.

  Report your findings in a straightforward manner with no conclusions or value judgements as to what the data might mean. Give a detailed report on the facts in a logical order including micrographs, graphs, tables or drawings that must be clearly labeled and captioned. Include a written description which describes the results illustrated in the graphs or figures. The narrative writing points out trends or inconsistencies, but should not include explanations or opinions. The text calls attention to what you consider to be significant findings that you will explain in the Discussion section.

  When describing results, use direct sentences with important subjects and active verbs. Avoid weak phrases such as “Fig. 1 shows that...”. Instead say, “Xylem was arranged in a circle around the pith (Fig. 1).” Pay attention to subject- verb match: What is wrong with the observation that “The Petri dish grew...”?

  When table or graphs are used, data should be presented in a form that easily shows the reader correlations or patterns that are important. Here are specific requirements for tables and figures:

  • Columns of numbers or words are TABLES; all other items (graphs, pictures, drawings, maps, etc.) are FIGURES.
  • Number tables and figures separately in the order in which they are mentioned in the text.
  • All tables and figures must have self-explanatory titles. The reader should be able to look at a table or figure title and know what was done in that part of the experiment.
  • Tables and figures should include units of measure (g, m, sec, etc.). All columns in a table should have legends; X- and Y-axes on a graph must have legends with units.
  • When a table or figure is included, there must be some reference to it in the text. For example, "The optimum temperature is 37o C (Fig. 1)." Such a signal refers the reader to that specific figure.

  DISCUSSION.

  Interpret the meaning of your results. Describe patterns or correlations that you think are important. Offer an explanation for your observations. If they turned out differently from what you had expected, give possible explanations.

  Explain how your findings are similar to or different from those of others. Compare your results with those in published works. Paraphrase the findings of others.

  Make conclusions about the meaning of the data; defend your conclusions. What were the answers to questions from the Introduction? Was your hypothesis supported? What are the main principles shown by your results?

  LITERATURE CITED.

  List in alphabetical order by author, all publications referred to anywhere in the text of the paper. Literature Cited includes only those references that were actually mentioned or cited in the paper. Other information that you may have read about the problem, but did not mention in the paper, is not included. That is why the section is called Literature Cited, not Bibliography.

  Any time you mention information that you have not obtained from your experiments or observations, you must refer to the source of that information. To cite a reference, in a sentence in which you have stated information from a reference, put the name(s) of the author(s) and the year in parentheses at the end of the statement. For example:

  Some birds are insectivorous and obtain all the water they need from the body fluids of insects (Bartholomew and Cade 1963).

  Information given to you by conversation is weak because no one can look it up. It is appropriate only for unpublished information. Such are not listed in Literature Cited, but as follows:

  Some birds never drink (R . Jones, personal communication).

  Details of punctuation, order of information, etc, in Literature Cited are complex. Here are examples of the format to use:

  Bartholomew, G. A., and T. J. Cade. 1963. The water economy of land birds. Auk 80:405-410

  Cade, T. J. 1959. Sea water and salt utilization by Savannah Sparrows. Physiol. Zool. 32:320-333

  Gubanich, A. A. 1977. Writing the scientific paper in the investigative lab. Amer. Biol. Teacher. 39:27-31.

  Humason, G. L. 1962. Animal Tissue Techniques. W. H. Freeman and Co., San Francisco, pp. 101-105

  Keeton, W. T., and J. L. Gould. 1993. Biological Science. W. W. Norton and Co., New York, pp. 207-208

  Checklist of critria used for grading lab reports

  Title

  Does the title explain the content of the paper?

  Summary

  Does the summary include the purpose, the methods, the results and the conclusion? Is it brief?

  Introduction

  Is there a clear statement of the problem?

  Is the background appropriate for an understanding of the problem?

  Is the background accurate?

  Is the problem or question important?

  Is a hypothesis given? Is the rationale for it explained?

  Materials and methods

  Is sufficient information given to permit duplication of the work? Is it concise?

  Results

  WRITTEN (PROSE) PART:

  Are results given concisely in a logical manner?

  Are trends or patterns pointed out?

  Are the results quantitative?

  Does the prose description refer to all tables and figures?

  Are results given for all procedures described in Materials and methods?

  No interpretations or opinions belong in the results, just data and descriptions.

  FIGURES

  Are tables and figures clearly labeled including figure legends?

  Do tables and figures have captions which describe precisely how the data were obtained?

  Do figures show what written part says they show?

  Discussion

  Are generalizations or conclusions derived from the results?

  Are they appropriate or consistent with the results?

  Are the observations explained?

  Does the discussion apply the results to the original question or hypothesis?

  Are the conclusions compared with those of earlier workers?

  Is the discussion clear and to the point?

  Literature Cited

  Is the format correct?

  Are all listed papers cited somewhere in the paper?

  Are all cited papers listed in this section?