Investigating Number of Stomata on a Leaf

Lab Design “Investigate the effect of a factor on the number of stomata of a leaf. ”

Research Question

How do differing leaf heights affect the number/density of stomata of a leaf? Hypothesis Stomata are pores, typically found under the leaf (lower epidermis), that control the gas exchange of transpiration, where water vapor leaves the plants, and carbon dioxide enters. I predict that the stomatal density on high leafs is higher than on low leafs. During photosynthesis the chloroplasts in the leaf cells synthesize ATP from ADP as a result of exposure to light, while oxygen is produced as a by-product of the photosynthetic reaction.

Carbon dioxide, which enters the plant through diffusion via the stomata, is needed for this process (photosynthesis) to occur. When the chloroplasts in the leafs cell is exposed to higher light intensities, more ATP is synthesized from ADP, while production of the by-product oxygen also increases. This increase in the rate of photosynthesis calls for more “fuel”, i. e. Carbon dioxide. So for a higher concentration of carbon dioxide to diffuse into the plant, the plant must grow a greater stomatal density (higher number of stomata).

This will create a larger surface area for carbon dioxide diffusion, the excretion of water vapor (transpiration) and the large amounts of oxygen being produced. As the higher leafs are exposed to higher light intensities I predict the stomatal density to be high. Lower leafs are exposed to lower light intensities due to, for example, shading by top leafs, and will so have a lower stomatal density than high leafs.

Variables Controlled

Type of plant- The type of plant that is going to be used will stay the same, i. e. controlled. The type of plant that is used for this experiment is called Quercus Ilex. Amount of leafs (10 ‘high’ leafs, 10 ‘low’ leafs)- the ensure fair testing the number of leaves tested from each variable will be the same. Apparatus used- Same set up each time. Microscope magnification (400x)- Magnification at which the number of stomata will be counted at is at a magnification of 400x.

Independent Variable

• Leaf Source

The leaf source regarding to the ‘high’ and ‘low’ leafs is the variable which will be changed to test the difference in number of stomata of the two variables.

• Distance between high/low leafs

The distance between the height at which ‘low’ and at which ‘high’ leaves were picked each time had to be of a minimum of 20cm to ensure plausible results. Lower epidermis of leaf used to count stomatal density- Because Quercus Ilex is a dicotyledonous plant, the number of stomata on the lower epidermis will be higher than on the upper epidermis. This is because dicotyledonous plants hold up their leaves horizontally, which directly illuminates the lower epidermis. So, to prevent water loss, fewer stomata will then be located on the upper epidermis.

Dependent Variable

Stomatal Density of high leafs Stomatal Density of low leafs Apparatus/Material 10 high leafs 10 low leafs Clear nail polish Slides Pincette Microscope Clear Tape Calculator Method Find a leaf source that has a significant height from which you will be collecting your leafs from throughout the entire experiment. Determine a low area, of little height from the ground, on the source from which you will pick 10 ‘low’ leafs. Repeat step 2, except that the area must be at an increased height distance of at least 20cm, to ensure a fair test and collection of ‘high’ leafs from a higher area than that of the ‘low’ leafs.

Choose a leaf of which the stomatal density is to be examined but don’t pick it off the plant. This is so that the plants photosynthetic process will not be disturbed which could lead to change in the leafs natural state and affect your results. Paint a layer of clear nail polish on the lower epidermis of the leaf and wait until it has dried. Use your tweezers to gently peel off the dried layer of nail polish. Gently peel the area of dried nail polish from the leaf completely. You should see a cloudy impression of leaf surface on the piece of tape. This is the leaf impression. Place the leaf impression to a clean slide.

Label the slide for identification if necessary. Focus the leaf impression under a microscope magnification of 40x until it is focused and observe the leaf impression. Find an area that is clean of thumbprints, away from the edge of impression, has no damaged areas or big leaf vein impressions in view. When focused, observe the impression under an increased microscope magnification of 100x and make sure it is focused. When focused, observe the impression under an increased microscope magnification of 400x, the magnification at which you will count the number of stomata, and focus.

Count the number of stomata you see in the field of view and record the number in a table under the relevant variable (‘high’ or ‘low’ leaf). To ensure a fair test, repeat steps 9-13 two times by choosing a new spot on the same leaf to focus on. Use the higher number of the 2 repeats to find the average later on. Repeat steps 1-14 ten times for the 10 high leafs and 10 low leafs.

Raw Data

How differing leaf heights affect the number/density of stomata of a leaf? One manipulation that was done to the raw data to help make it more useful for interpretation was the rounding off.

Calculations Difference in mean > 46. – 26. 2 = 20. 1 Difference in S. D. > 6. 993 – 2. 3 = 4. 693 Because the standard deviations are much less than the difference in the mean number of stomata, it is very likely that the difference in the mean number of stomata between High Leafs and Low Leafs is significant.

T-TEST Null hypothesis: The number of stomata on high leafs and low leafs are not different. The differences in the data sets are the result of chance variation only and they are not really different. Mean of # of stomata on High Leafs: 46. 3 Mean of # of stomata on Low Leafs: 26. 2 t=8. 63 Degrees of freedom= (10+10)–2= 18 Critical value for t=2. 101 (P= 0. 05)