Materials Included In Kit

Additional Materials Required

Prelab Preparation

1. Bean plants should be planted about two weeks before the start of this activity.
2. Fill each of the watering trays with soil.
3. Place rows of bean seeds in the soil approximately 1" deep in both trays. Plant an excess of bean seeds in case some of the seeds fail to germinate.
4. Water and cover the planting tray with plastic wrap to keep the seeds warm and speed up germination.
5. Place the trays under a grow light or in a sunny location. Remove the plastic wrap after the seeds have germinated.
6. Keep the soil moist over the two-week growing period.

Safety Precautions

Although the materials in this lab activity are nonhazardous, follow normal safety precautions. Remind students to wash their hands thoroughly with soap and water before leaving the laboratory.

Disposal

Please consult your current Flinn Scientific Catalog/Reference Manual for general guidelines and specific procedures, and review all federal, state and local regulations that may apply, before proceeding. All materials may be disposed of in the regular trash according to Flinn Suggested Disposal Method #26a. 

Teacher Tips

• Enough materials are provided in this kit for 30 students working in pairs or for 15 groups of students. Both parts of this laboratory activity can reasonably be completed in one 50-minute class period. The observations for Part II should be recorded the next day or class period.
• The bean seeds should be planted roughly two weeks before performing this activity. See the Prelab Preparation section for more information.
• Have students share the 5 g foil packs of petroleum jelly. Vaseline^® may also be used. Have students only make one leaf impression on each bean leaf.
• Depending on the level of your students, you may wish to have them determine the total number of stomata per square millimeter. Do so by using the following equation:
  {10816_Tips_Equation_1}
  Assuming the 40X objective has a field diameter of 0.45 mm (r = 0.23 mm), the field of view would be calculated to be:
  
  Surface area of the field of view = π x 0.23 mm²

  = 0.17 mm²

  Equation 2 can be used to determine the number of stomata in 1 mm².
  {10816_Tips_Equation_2}
  For example, if 5 stomata were found in the 0.17 mm² field of view, 29 stomata would be found in 1 mm².
  {10816_Tips_Equation_3}X = 29 stomata
• Once the technique of “lifting” stomata prints has been mastered, students can generate questions and make predictions about variables that might affect stomata density. Students can then design their own experiments and collect data to determine the validity of their hypotheses. A few possible extensions are as follows:
  • How does the number of stomata in the upper and lower surfaces of leaves compare?
  • How do stomata densities vary from species to species?
  • Do the densities vary in the same species growing in different areas?
• The plastic test tubes may be taped directly to a window or placed in test tube racks for Part II. If the test tubes are taped to the window, make sure this activity is done during a fairly warm time of the year.

Introduction

What are stomata and how do they regulate the water balance in a plant? Perform the following activities and find out.

Concepts

• Stoma
• Guard cells
• Transpiration

Background

Plant tissue, just like animal tissue, is composed of specialized cells to perform specific functions. Plants have an epidermis layer, an outer skin-like layer, just like animals. Animal skin contains specialized “holes” or pores that regulate different body functions. Plant epidermis likewise has pores. A single pore in plant epidermis is called a stoma (plural—stomata). The number, location, and density of these pores are different for different plants—depending on their “niche” roles in different habitats. Stomata are most numerous on the leaves of plants. They occur on both the upper and lower epidermis of the leaves in some species (alfalfa, corn), exclusively on the upper epidermis in other plants (water lily), and are absent altogether on submerged leaves of aquatic plants. Stomata are very numerous, ranging from about 1,000 to more than 1.2 million per square centimeter of plant tissue. An average-sized sunflower leaf, for example, has about 2 million stomata on its lower epidermis.

Each stoma is bordered by two sausage-shaped cells that are usually smaller than surrounding epidermal cells. These small cells are called guard cells and, unlike other cells in the epidermis, they also contain chloroplasts (see Figure 1).

The photosynthesis that takes place in the guard cells aids in the functioning of these cells (i.e., the opening and closing of the stomata openings). This regulated opening and closing of the pores allows carbon dioxide (CO₂) and oxygen (O₂) to be exchanged between the interior of the leaf and the outside atmosphere. The opening and closing of the stomata also helps regulate the water balance inside the plant—water can more easily escape when the stomata are open. The process of plants losing water through the stomata is known as transpiration. Many environmental conditions influence the opening and closing of the stomata and thus affect the rate of transpiration. Temperature, light intensity, air currents, humidity and the nature of the plant all influence the guard cells to open or close.

It is the unique structure of the guard cells that allows the opening and closing to occur. Internal microfibrils and thicker inner walls of the guard cells cause these guard cells to “bulge” when osmotic pressure builds up inside them. When the water content of the guard cells is high the stoma is open and when the water content is low the stoma is closed.

Experiment Overview

The purpose of this experiment is to observe the properties of stomata under a microscope, count the average number of stomata in a plant leaf, and investigate the effect of different factors on the rate of transpiration.

Materials

Safety Precautions

Although the materials in this lab activity are nonhazardous, follow all normal laboratory safety guidelines. Wash hands thoroughly with soap and water before leaving the laboratory.

Procedure

Part I. Viewing Stomata

1. Obtain a leaf from one of the bean plants.
2. Paint a thick patch of clear nail polish on the lower surface of the leaf. Make the patch at least one square centimeter in size.
3. Allow the nail polish to dry completely.
4. Tape a piece of clear cellophane tape to the dried nail polish patch.
5. Gently peel the nail polish patch from the leaf by pulling on a corner of the tape and “peeling” the fingernail polish off the leaf. This is the leaf impression that will be examined.
6. Tape your peeled impression to a very clean microscope slide. Use scissors to trim away any excess tape.
7. Examine the leaf impression under a compound microscope at 400X magnification. Draw a picture of an individual stoma on the Stomata and Transpiration Worksheet.
8. Search for areas on the leaf where there are numerous stomata, and where there is no dirt, thumbprints, damaged areas, or large leaf veins.
9. Count all the stomata in one microscope field. Record the number on the Stomata and Transpiration Rates Worksheet.
10. Repeat counts for at least two other distinct microscope fields at the same magnification. Record all the counts on the Stomata and Transpiration Rates Worksheet.
11. Determine the average number of stomata using the three distinct microscope field counts. Record the average on the Stomata and Transpiration Rates Worksheet.
12. Answer the questions for Part I on the Stomata and Transpiration Rates Worksheet.

Part II. Transpiration Rate and Stomata

1. Place four test tubes in a rack.
2. Select four bean plants that are similar in height and have similar sized leaves.
3. Using scissors, cut the stems of the four bean plants at their base (where the stem meets the soil).
4. Put the plant cuttings into the test tubes (see Figure 2).
   {10816_Procedure_Figure_2}
5. Fill each test tube to the 12-mL mark on the tube with water.
6. Carefully cut off one of the leaves (the seedlings should have two leaves) of one of the plants—this will be plant 1.
7. Carefully remove both leaves of another plant. This will be plant 2
8. A third plant will not have any leaves removed. This will be plant 3.
9. On the fourth plant, smear petroleum jelly on the upper and lower sides of both leaves until completely covered. This will be plant 4.
10. Record the initial water volumes (in mL) for plants 1, 2, 3 and 4 in the Stomata and Transpiration Rates Worksheet.
11. Predict what will happen to the water level in each plant test tube setup. Record these predictions in the Stomata and Transpiration Rates Worksheet.
12. Place the plants under a grow lamp or next to a window in an area where they will remain undisturbed according to your teacher.
13. The next class period, observe the new water level of each test tube, in mL. Record all of the final volumes in the Stomata and Transpiration Rates Worksheet.
14. Answer the questions for Part II on the Stomata and Transpiration Rates Worksheet.
15. Consult your instructor for appropriate disposal procedures.

Student Worksheet PDF

10816_Student1.pdf