PLANT PHYSIOLOGY LAB (Last one---2+
weeks worth)
Tropisms, Nutrition, and
Growth Regulators
OBJECTIVES
By
the end of this exercise you should be able to:
1.
Define the terms phototropism and gravitropism.
3.
Explain the modes of action of auxin and gibberellic acid.
Plants
respond to a variety of environmental stimuli such as light, gravity, and
require several nutrients such as calcium and potassium. Plant growth and
development are controlled by internal chemical signals called growth
regulators. Auxin and gibberellic
acid are examples of growth regulators in plants.
In
this exercise you will study some common physiological responses of plants.
Remember that each of these responses is part of an overall design for the
survival and reproduction of plants in varied environments.
PLANT
TROPISMS
A
tropism is a movement in response to an external stimulus where the direction
of movement is determined by the direction of the most intense stimulus. Two
tropisms you will study in today's lab are phototropism, which is directed
growth in response to light, and gravitropism, which
is growth in response to gravity.
Phototropism
PROCEDURE:
OBSERVE PHOTOTROPISM
1.
Obtain from your lab instructor some 10-to-14-day-old radish (Raphanus) seedlings. These seeds have been grown in
diffuse, overhead light.
2.
At the beginning of the lab, place your seedlings approximately 25 cm from a
100 watt light so that light strikes the shoots at a right angle.
3.
Use a protractor to measure curvature of the seedlings every 30 minutes for 2 hours.
4.
Record your results in the following chart.
|
Time
(h) |
Curvature
(degrees) |
|
0.5 |
|
|
1 |
|
|
1.5 |
|
|
2 |
|
QUESTION
1
a.
Which direction did the seedlings curve?__________________________________________________
b.
Is this curvature positive or negative phototropism? __________________________________________________
c.
What is the adaptive significance of phototropism? __________________________________________________
d.
Would you expect roots to react similarly to light? Why or why not? _____________________________________
QUESTION
2
a.
Which roots grew down? __________________________________________________
b. Which one(s) didn't? __________________________________________________
c.
What conclusion about the part of the root that perceives gravity do you draw
from these data?________________
d.
Where in roots does the differential growth occur that produces gravitropism? ________________
e.
Are roots positively or negatively gravitropic? ________________
f.
What is the adaptive significance of gravitropism? ________________
QUESTION
3
a.
How are the stems now oriented? __________________________________________________
b.
Why is this response important in plants? __________________________________________________
Gravitropism
Yesterday,
onions having roots approximately 1 cm long were placed in a glass beaker.
Seedlings labelled "H" had their root
oriented horizontally, while those labelled
"D" had their root pointing down. The terminal 3-4 mm was removed
from roots of seedlings also labelled with an
asterisk (*). Examine the experimental set-up, and record your results in the
following chart.
|
Treatment
or orientation |
Direction
of Growth |
|
Intact
roots oriented vertically (V) |
|
|
Intact
roots oriented horizontally (H) |
|
|
De-tipped
roots oriented vertically (V*) |
|
|
De-tipped
roots oriented horizontally (H*) |
|
Examine
the Coleus that yesterday were inverted or oriented vertically or horizontally.
Record your observations in the following chart.
|
Orientation |
Response |
|
Vertical |
|
|
Horizontal |
|
|
Inverted |
|
Gibberellic Acid (GA)
Dwarfism
often results from a plant's inability to synthesize active forms of gibberellic acid (GA), another plant growth-regulator. Gibberellic acid promotes stem elongation and mobilizes
enzymes during seed germination. Examine the four trays of corn plants that
were treated in the following ways:
Tray
1:
Tray
2: Dwarf plants-untreated
Tray
3:
|
Plant-treatment |
Observations |
|
Normal-untreated |
|
|
Dwarf-untreated |
|
|
Normal-treated |
|
|
Dwarf-treated |
|
b.
Did applying GA to normal plants have any effect? If so, what? _________________________________
There
are plant movements other than tropisms, but they will not be considered in
this exercise. Consult the text, lecture notes, and references if additional
information is desired.
A.
Negative Geotropism of Stems
·
Place
a potted sunflower plant 15 to 20 cm tall in a horizontal position on the
laboratory table.
·
Carefully
measure the shortest distance from the tip of the stem to the surface of the
table. ______________
·
Repeat
this measurement at fifteen minute intervals. Also note any bending of the stem
and the location of the region of curvature.
·
Compare
this with the location of the region of curvature in a plant which has been
lying in a horizontal position for twenty-four hours. What differences are
there between the two plants? __________________________________________
·
Suggest
a possible explanation for these differences. __________________________________________________
·
Before
the close of the period sketch the plant, showing the curvature as it then is.
Indicate by dotted lines the original position of the stem.
B. Geotropism of Roots and Stems
Seeds
of mustard, radish, or cress will be germinated in the dark on moist filter
paper in Petri dishes, the covers sealed in place with gummed paper or Scotch
tape. Each dish has been kept on its edge by use of a clamp or other means.
Note that the seedlings have developed definite roots and stems and that they are growing in a vertical position. Mark on the
glass with wax crayon the position of the seedlings, then
rotate the petri dish through ninety degrees so the
seedlings are now horizontal. Clamp in place and observe at the end of the
laboratory period, at the beginning of the next laboratory period.
·
What
changes in the position of the seedlings are evident? __________________________________
·
What
typical responses to gavity are exhibited by the
roots? __________________________________
·
By
the stems? __________________________________________________
·
Make
a sketch of the seedlings, indicating the position before (by dotted lines) and
after (by solid lines) their position was shifted.
C. Phototropism
Pots
of oat, radish, or wheat seedlings will be grown until they are 4 to 5 cm tall.
Keep overnight in darkness before beginning the experiment.
Still
keeping the seedlings in darkness, place one of the pots in a light-tight box
which has a 7-mm opening on one side.
Expose to daylight so light is admitted only
through the opening.
Prepare a similar setup as a control but
without the opening in the light-tight box.
At
the end of two hours remove the boxes and observe results. Allow another set of
seedlings to remain undisturbed until the next laboratory period. What has
happened? __________________________________________________
Sketch to show position of seedlings before
and after exposure to one-sided illumination.
Comment briefly on the tropistic
movements of plants and their parts as they occur in nature.
What advantage are these movements to the
plant? __________________________________________________
No. 3. Record results as indicated in the above directions.
QUESTIONS
1.
What causes a stem to bend following a stimulus such as light or gravity? __________________________________________________
2.
How does the characteristic response of roots and stems to hormones apparently
determine the vertical growth habit of a typical plant? __________________________________________________
3.
What is a tropism? __________________________________________________
4.
What is meant by positive and negative responses? __________________________________________________
5.
List a number of forces or stimuli that cause tropisms and give the
characteristic response of the plant to each. __________________________________________________
6.
What part do growth substances play in causing tropisms? __________________________________________________
7.
Describe a number of tropistic responses of plants as
they are found in nature and list the possible advantages of each to the plant.
__________________________________________________
D. Gravitropic
and Phototropic Curvature in Coleus
Materials
(on demonstration) Coleus plant placed on its side Coleus
plant in unilateral light Coleus plant in upright position
Introduction
In
this lab study, you will investigate the growth of the stem in response to two
environmental stimuli, gravity and light. Gravitropism
(or geotropism) is the response of a plant to gravity. Phototropism is the
response of a plant to light. Three Coleus
blumei plants are on demonstration in the lab
room. One was placed on its side several days ago. Another has been growing in
unilateral light for several days. The third, the control, was left undisturbed
in the greenhouse until lab time.
Procedure
1.
Study gravitropic curvature in Coleus blumei.
a.
Carry your lab notebook to the demonstration area and observe the Coleus plant
placed on its side.
b.
Examine the plant, noting the appearance of different regions of the stems and
roots (if visible). What part of each stem has curved? To
what degree? __________________________________________________
c.
Compare this plant with the control plant left in an upright position. _______________________________________
d.
Describe your observations ___________________________________________
2.
Study phototropic curvature in Coleus blumei. __________________________________________________
a.
Examine the plant growing in unilateral light, noting the appearance of
different regions of the stems.
b.
Compare the plant to the control plant, which has received light from all
directions in the greenhouse.
c.
Record your observations and answer the questions.
E. Bending Response from Auxins
Select
a vigorous potted tomato plant 25 to 50 cm tall. By means of a ring stand and
clamp, fasten a meter stick in a vertical position near the plant. With a
thread, attach a finely drawn (hairlike ) piece of glass rod or thin capillary tubing to a petiole
so the end of the rod extends across the meter stick. Record the reading on the
meter stick and
then apply a liberal smear of three percent indolebutyric
acid or indoleacetic acid in lanolin to the upper
surface of the petiole. Record time of hormone application.
Set up a control experiment using pure lanolin only. Observe any bending of the
petiole by checking the movement of the glass rod along the meter stick. How
many centimeters does the glass rod move in the treated and control plants,
respectively, in thirty minutes? one hour? two hours? overnight? Why does the
petiole bend downward when the auxin smear is applied
to the upper surface? Is this bending a true growth response? Why?
F. Epinasty
Observe
the experiment set up by the instructor to demonstrate the effect on plants of
ethylene gas released from ripe fruit. This is shown by placing a bell jar over
a tomato plant together with several pieces of ripe apple, banana, or other
fruit. The petiole of a rhubarb leaf may also be used. The tomato plant is very
sensitive to ethylene gas; one part of gas in two million parts of air is
enough to induce the characteristic response. The experiment should remain in
the dark for sixteen to twenty-four hours. Compare this setup with a control
without the fruit. Leafy cuttings from a tomato plant (kept in a tumbler of
water) will exhibit the same response.
No. 1. Describe results and make quick sketches to show the effect
of ethylene gas from ripe fruit on the tomato plant. Suggest a possible
explanation of this behavior.
G. Gibberellic
Acid
Select
four dwarf bean plants 5-10 days old that are the same size and height.
Determine and record the height of each by measuring from the soil line to the
stem tip. Note especially the length of the internodes.
Spray
two experimental plants until the leaves are wet (almost dripping) with a one
part per million (ppm) solution of gibberellic acid in distilled water. Label and store in the
greenhouse.
Similarly
spray two control plants with distilled water until the leaves are wet. Spray,
label, and store in the greenhouse some distance away from the experimental
plants.
Measure
and record the height of each plant five days after spraying. Are there any
differences between the gibberellic acid-treated
plants and the controls? Repeat the above observation 10 days
after the original treatment and at succeeding intervals of 5 days each
thereafter, or until no further changes are noted. Are any differences
more evident after 10 days? 20 days? 30 days? Are all the plants still the same
height? Are the internodes the
same length and number in both control and experimental plants?
Write a short paragraph giving and explaining the results of this experiment.
D.
Root Formation
H. ROOTING OF CUTTINGS
Treat
the base of 8-to-10-cm geranium, Coleus, Bryophyllum,
or tomato cuttings with a hormone powder or smears of 3 percent indolebutyric acid or alpha-naphthalene acetic acid as
directed by the instructor. Place cuttings in a flat of sand at a depth of
about 5 cm. Plant another group of similar cuttings but omit the hormone
treatment. Remove both groups from the sand in ten to twenty days and record
the percentage of rooting in each case. Sketch results.
Is there any difference in the rooting of treated and untreated cuttings?
I. ROOTING OF LEAVES
Apply
a lanolin smear of three percent indolebutyric acid
or of alpha-naphthalene acetic acid near the tip of several Coleus leaves (do
not remove from plant). Observe in ten to twenty days. Do roots form on the
bottom or top of the petiole? Sketch results, showing any
roots and indicating changes in the position or form of the leaves.
J . Seedless (Parthenocarpic) Fruits
Pollen
and pollen tubes contain a natural growth substance which is responsible for
fruit development, chiefly by stimulating the growth of the ovary. If a
synthetic hormone possessing similar properties is applied to the ovary, fruit
formation also occurs, but such fruits will have no seeds. Synthetic hormones
may be applied to a flower in lanolin smears, as sprays or as a gas.
Considering the usual sequence of events in pollination and fertilization, can
you explain why these artificially induced fruits are always seedless? Why may
a seedless fruit be said to contain abortive ovules? Name several seedless
fruits sold commercially. How were they developed? How are seedless fruits
propagated?
K. HORMONE SPRAYS
List advantages of sprays over lanolin smears for this type of
work.
L. Weed Control
A
number of synthetic growth substances kill plants when applied to them. Because
of this property these compounds may be used as herbicides to kill or control
undesirable plants such as weeds.
Characteristics
of these compounds and their effects on plants may be briefly listed: (1) they
are not caustic, (2) they are active in small concentrations, (3) the effects
often appear some distance from the part where applied, (4) leaf growth and
development of buds are reduced, (5) epinasty, contortion
of petioles and stems, and abnormal development of plant parts occur, (6)
metabolism is seriously deranged, (7) reserve foods are exhausted, and (8) they
are extremely selective.
A
herbicide commonly used in the
of
its derivatives. This chemical may be applied as a salt, an amide, or as an
ester. In general, the salt is applied to susceptible (easy to kill) plants;
the esters to woody or resistant (hard to kill) perennials. A strong solution
of 2, 4-D will kill practically any plant, however, concentrations of one
thousand to two thousand parts of 2, 4-D per million parts (ppm)
of water will kill most broad-leaved (Dicot ) plants
but usually causes little or no injury to narrow-leaved (Monocot) plants.
General directions for the use of 2, 4-D cannot be given with safety since the
amount applied and the type of 2, 4-D compound used is dependent on greatly
varying local conditions and the species of weeds to be killed.
M. EFFECTS ON BROAD- AND NARROW-LEAVED PLANTS
Corn
and beans have been planted together at the same time in 15-to-20-cm pots. When
the corn plants are about 15 cm tall the beans are usually large enough for the
experiment. Spray two pots of corn and beans with a 1500 ppm
solution of the salt of 2, 4-D (Note: The actual amount of 2, 4-D salt in
commercial products may vary considerably; however, if the directions given on
the container are carefully followed, the resulting solution will contain
approximately 1000 to 1500 ppm of 2, 4-D). Keep at
least one pot unsprayed as a control. Treated plants must be sprayed outdoors
or in a room away from the controls so that no trace of the chemical comes into
contact with them. Examine all plants at three-day intervals for two weeks or longer
and make quick sketches and brief notes at each observation. What happens to
the bean plants? Does the killing appear to begin with the leaves and progress
down the stem into the roots or do the symptoms appear in reverse order? Does
the appearance of the bean plants indicate that a growth substance may be
exerting its influence? State reasons for your answer.
Do the roots of the bean eventually die? Are the corn plants injured in any way
by the chemical? __________________________________________
_______________________________________________________________________________________________
.
At the conclusion of the experiment, write a short summarizing statement
accompanied by sketches to record results of this work.
QUESTIONS
1.
What are plant hormones? What are their characteristics and properties? ___________________________________
2.
How do they act in causing bending of plant stems? __________________________________________
3.
Are plant hormones used in the same part of the plant in which they are formed?
_______________________________
4.
In what parts of the plant are hormones produced? __________________________________________
5.
What is the commercial importance of plant hormones? __________________________________________
6.
How recent is most of the work on plant hormones? __________________________________________
7.
What is lanolin? __________________________________________
8.
Explain how seedless fruits are formed by the application of synthetic hormones
to flower parts. Do seedless fruits occur naturally? __________________________________________
9.
Compare seedless fruits with normally developed seeded fruits. Consider color,
firmness, flavor, and food value. __________________________________________
10.
What are herbicides? Name several methods of weed control not involving the use
of 2, 4-D. __________________________________________
11.
List symptoms and give the order of their occurrence of a plant treated with a
toxic application of 2, 4-D. __________________________________________
12.
List precautions to be observed in the use of 2, 4-D. Suggest reasons for
variable results following the careless use of this herbicide. __________________________________________
REFERENCES
BIDWELL,
R. G. S. 1974. Plant physiology.
DEVLIN,
R. M. 1973. Plant physiology.
MEYER,
B.; ANDERSON, D. B.; BOHNING, R. H.; and
FRATIANNE,
D. C. 1973. Introduction to plant physiology. 2d ed.
STEWARD, F. C., and KRIKORIAN, A. D. 1971. Plants,
chemicals and growth.
WAREING, P. F., and PHILLIPS,
Additional Investigations
of Auxins
Materials
auxin solutions in various corn
and bean seedlings in pots or flats
concentrations lamps
auxin in lanolin paste toothpicks
lanolin with no auxin protractor
scissors spray bottles
Coleus
plants cotton-tipped applicators
glass containers for planting aluminum
foil Brassica rapa seedlings in quads
Introduction
Having
made observations of gravitropism and phototropism in
the preceding lab study, discuss with your team members ways to use Coleus, Brassica, or corn or bean seedlings to further investigate
these phenomena. If you choose to carry out your independent investigation with
this system, the questions provided in the following Procedure section will be
appropriate for your study. Using the materials available,
design a simple experiment to investigate the role of auxin
in plant growth or factors involved in phototropism and gravitropism.
Procedure
1.
Collaborating with your research team, read the following questions, check your
text and other sources for supporting information, and choose a question to
investigate, using this list or your own imagination.
a.
If only some wavelengths stimulate phototropic response, which ones do and
which do not?
b.
If the apical meristem is removed, will plants
respond to unilateral light?
c.
If the tip of the root is removed, will roots respond to gravity? (Seedlings
can be planted close to the wall in glass containers so that root growth can be
viewed.)
d.
Can these tropisms be altered by applying auxin paste
to the plant?
e.
What will happen if the tips of the plants (root or stem) are covered with
aluminum foil?
f.
How else does auxin affect plants? How does auxin affect apical dominance? Can auxin
be used as an herbicide? (In what concentrations? What
is the effect on plants?) What concentration of auxin
produces the largest roots on cut stems? (What horticultural application would
this have?)
g.
Will seedlings growing in the dark respond to auxin
applied to the side of the stem? At all locations on the
stem?
2. Design your experiment,
proposing hypotheses, making predictions,and
determining procedures
Plant
Growth
Have
each student team record its final results on the board or on an overhead projector acetate. Then students can calculate
mean values for the entire class.
Results
1.
Determine and record the mean height of plants in each category in Table 20.1.
2.
Using the mean height for each category of plants,
calculate the percentage difference in the mean height of treated normal plants
and control normal plants. Then calculate the percentage difference in the mean
height of treated dwarf plants and control dwarf plants. Use the given formula
for your calculations:
Normal
% difference = treated - control x 100 = %
control
Dwarf
% difference = treated - control x 100 = %
control
3.
Record your data for the average percentage difference in the mean values for
both normal and dwarf plants from Table 20.1 on the class master sheet. Then
calculate the average percentage differences for the entire class.
Your Data Class Data
Average
% difference: normal
dwarf
Discussion
1.
How do values for percentage difference compare for dwarf versus normal treated
and untreated plants?
The
percentage difference in height is much greater between dwarf treated and dwarf
untreated than between normal treated and normal untreated. The percentage
difference in height of normal treated and normal untreated should approach
zero.
2.
What is the action of gibberellins? Discuss your results with your group, and
consult your text or other references in the laboratory.
The
dwarf condition is due to a mutation that results in no gibberellin
synthesis. Adding gibberellins reverses this phenotype, causing the internodes
to elongate and the plants to grow to normal phenotype. Adding gibberellins to
plants that are normally tall has little effect on size. Apparently, these
plants have adequate gibberellins present to attain maximum height.
Lab Study B. Additional
Investigations of Gibberellins
Materials—use
results from your hormone growth seeds
normal and dwarf Brassica rapa seedlings solutions of gibberellin
normal and dwarf corn seedlings dropper
bottles
normal and dwarf pea seedlings sprayers
(Little
Marvel peas, Pisum sativum) cotton-tipped applicators
Introduction
Having
seen the effect of gibberellins on the growth of normal and dwarf corn
seedlings in the preceding lab study, discuss with your team members possible
questions for further study of this group of hormones. If you choose to carry
out your independent investigation with this system, the questions provided in
the following Procedure section will be appropriate for your study. Using the materials available, design a simple experiment to
investigate the actions of gibberellins in plant growth or seed germination.
Procedure
1.
Collaborating with your research team, read the
following questions,
check your text and other sources for supporting information, and
choose a question to investigate, using this list or your own
imagination.
a.
Plant scientists have discovered a mutant strain of Brassica
rapa in When purchasing
dwarf seeds, which plants are dwarf. In these plants, the internodes do not
elongate- order rosette seeds. gate, and plants
consist of a cluster of leaves spreading close to the soil. Flowers cluster
close to the leaves. What could be the cause of this phenotype?
b.
Would other plant hormones produce the same response in dwarf corn seedlings as
do gibberellins?
c.
In the demonstration experiment, the gibberellin
solution was sprayed on all parts of the plant. If the gibberellin
solution were added only to specific regions, such as the roots or apical meristem, would the effect be the same?
d.
Would the results in the corn experiment differ with different concentrations
of gibberellin solution?
e.
What effect do gibberellins have on seed germination?
f.
What effect do gibberellins have on root growth on cut stems?
g.
Is the dwarf condition seen in certain strains of peas (Little Marvel peas) due
to the lack of gibberellins?
2.
Design your experiment, proposing hypotheses, making predictions, and
determining procedures as instructed in Exercise 20.4.
Plant Growth Substances(Phytohormones)
Plant
Materials
Tomato
plants 20 to 50 cm tall (10-12 weeks old and in flower), ripe apple, banana, or
rhubarb stalk. Coleus, geranium, or tomato cuttings 7.6 to 10
cm tall. Corn and bean plants 15 to 20 cm tall grown together in sand
flats. Sunflower plants 20 to 45 cm tall and geranium plants 10 to 15 cm tall
grown in sand flats. Potted Coleus plant 20 to 25 cm tall.
Dwarf bean plants 5 to 10 days old.
Equipment
Ring
stands and clamps, meter sticks, strong thread, capillary tubing, fine glass
rod drawn out to a long hairlike thread, large bell
jars and glass plates, flat-ended toothpicks, small paper tags, India ink and
fine pen, melted paraffin. Hand atomizers, small thin spatulas, sharp
laboratory scissors (small), single-edged safety razor blades, curved sharp pointed
forceps.
Reagents
Three percent indole-acetic acid,
alpha-napthalene acetic acid, or indole-butyric
acid in anhydrous lanolin. Commercial hormone powder (Rootone or similar). A 1500 ppm
solution of the salt of 2, 4-D, 0.02 percent aqueous solution of beta-napth-oxyacetic acid, gibberellic
acid (1 ppm ).
Introduction
The
fact that phytohormones, also known as plant growth
regulators or growth regulators, are now an integral part of the knowledge
about plants, makes it desirable to know by
experiments some of the ways these compounds affect plants. Phytohormones
may be briefly characterized as substances of a definite chemical composition
which are synthesized by plant tissues and function as regulators
necessary for normal growth and development. They have a
definite point of origin (synthesis) in the plant but often exert their
specific effects in a part of the plant some distance removed from their point
of origin. Only very small amounts are needed to produce, marked physiological
effects. It has been estimated that hormones exist naturally in living plant
tissues in an effective concentration of only one part per million. Synthetic
hormones may be applied in effective concentrations of four to five parts per
million. Growth regulators may be broadly classed as auxins
(affecting cell elongation), kinins (influencing cell
division), gibberellins (stimulating elongation of stems and enlargement of
leaves), and inhibitors (a great variety of substances which inhibit growth).
Growth movements of plants are one common expression of hormone action.
Practical applications of hormones are made in horticulture and related
sciences in the rootings of cuttings, the production
of seedless fruits, the prevention of preharvest
apple drop, and as herbicides. The intelligent use of hormones is a standard
procedure for many horticultural projects, and some phases of this science have
been revolutionized by the use of hormones.