The Effects of Oil Contamination on Root Growth of Arabidopsis
Matt Steelman, Josh Fester, Tinus Van Wyk, and Nicole Sowers
Wofford College, South Carolina
Introduction
Some Arabidopsis ecotypes such as those from Ireland and Bulgaria
grow on roadsides where petroleum runoff could affect their growth.
In an experiment using phenanthrene as the pollutant, Arabidopsis
plants were shown to be negatively affected 1. Our purpose is to
investigate to what extent, if any, the root growth of Arabidopsis is
affected by contamination with motor oil. In this experiment, we used
ecotype Columbia because it is the most commonly used ecotype in
laboratory studies. Also, we chose our concentrations of motor oil in
agar with the desire to avoid killing the plant while providing adequate
range that would produce an observable response.
Results
Discussion
Germination
Germination was unaffected by oil dissolved in the agar
(p=0.667, χ2 with 6 degrees of freedom = 4.08, data not
shown.)
Root Growth
Firstly, germination was not affected by motor oil in agar, suggesting
that environment is less important than conditions inside the seed for
germination. Secondly, our results support the hypothesis that an
increased concentration of motor oil in agar stunts root growth.
Although there was no different in root growth from 0% oil to 0.01%
oil, but at the higher concentrations (0.1% and 1%) root growth was
significantly reduced compared to control. In general, root growth is
negatively affected by increased concentrations of motor oil.
Implications of this Experiment
Hypothesis
The addition of motor oil into the agar for Arabidopsis thaliana
ecotype Columbia will significantly stunt the growth of the root.
Methods
At the lowest concentration of oil (0.01%) root growth was
not significantly different from the control group. However,
root growth was decreased at the higher concentrations of
oil in agar (p=0.001, Analysis of Variance FdF= 3, 70 = 6.054.)
Root Growth and Oil Concentration
The results of this experiment show that while plants can tolerate low
levels of motor oil pollution, increased concentrations are detrimental to
root growth. It would be unwise to assume that all seedlings respond to
oil pollution in the same way as Arabidopis thalania ecotype Columbia.
It is likely that other ecotypes or other species of plants have greater or
lesser sensitivity to oil in the soil.
Future Experiments
Eight conventional Petri dishes containing 0.8% agar
Castrol motor oil (5W30 )
Eighty Arabidopsis (ecotype- Columbia) seeds
Image J Software
Fluorescent grow lights
We recommend that future experiments find a more effective way to
measure the root such as removing them from the agar. Also, the
experimenter must make sure that light is evenly distributed to each
agar plate. Another issue that should be addressed is clumping of motor
oil in the agar which may allow roots to avoid the pollution by growing
in the agar between the droplets of oil. Finally, effects of the motor oil
on root growth in agar may be different than those in soil, so perhaps
soil should be used in a follow-up experiment.
Standard agar was mixed with motor oil to obtain 0%, 0.01%, 0.1%,
and 1% dilutions. The agar containing motor oil required extensive
mixing to disperse the oil before pouring on to slanted petri dishes. We
then planted 10 seeds of Arabidopsis in each of eight Petri dishes; 2
dishes for each concentration . Seeds were placed at the upper edge of
the agar on each of the plates. Agar was used rather than soil so that we
could measure the roots without removing the plant from its substrate.
The dishes were sealed and vertically placed beneath fluorescent grow
lights at a distance of 33cm throughout the experiment. The two dishes
with 0% motor oil in agar served as the control group, while the other
six had the concentrations of the petroleum product. Each of the plates
had equal photoperiods (16 hours of light and 8 hours of dark) 33 cm
below a fluorescent light. Once a week for two weeks, we measured the
length of the roots using Image J software (see picture below) from
scans of each plate. We were able to accurately see and measure roots
and shoots for individually identified seedlings. Only root measurement
data are presented here..
References
1) Lui, Hong. “An oxidative stress response to polycyclic aromatic
hydrocarbon exposure is rapid and complex in Arabidopsis
thaliana.” Plant Science: Vol. 176 Issue 3 March 2009 pg. 375-382.
Acknowledgements
Bars are 95% confidence intervals.
Special thanks to GR Davis and C. Abercrombie for their assistance
with statistical analysis and answering of questions. Also, thanks to
A. Steadman and T. Player for their comments and suggestions.
Effects of LED and Fluorescent Lights on Root Growth of Arabidopsis
Kevin O’Quinn, Mesha Arant, Jordan Ball, and T.J. White
Department of Biology, Wofford College, Spartanburg, SC 29303
Introduction
Different types of lights have been used to increase
the growth of various plants. Our experiment tested
whether LED or fluorescent lights produce plants
with longer root length and higher germination rates.
Fluorescent lights, which emit heat, consume more
energy, and emit a more intense light are typically
used in laboratories. LED lights, which consume
10% less energy than fluorescent lights, can last for
many more years that fluorescent lights, but are more
expensive. Are the LED lights really worth the cost?
We performed an root growth experiment to find out.
Germination Rate
The germination rate, not shown, was not statistically different for seeds under fluorescent
and LED light (Chi-square (dF=1)=0.137; p=.712).
Root Length of Plants under Fluorescent and LED lights
1. Germination was not affected by light source.
2. Root growth was greater under fluorescent
lights.
Our results did not support our hypotheses.
Discussion
Hypothesis
1. Germination rates will be higher in plants grown
under the LED lights.
2. Root length will be greater in plants grown under
the LED lights.
Materials and Methods
Materials
•48 Arabidopsis seeds(Ecotype: Columbia)
•Petri dishes with 0.8% Agar
•Fluorescent and LED Growing Lights
Methods
•0.8% Agar was placed into 6 slanted Petri dishes.
•Eight seeds were placed in each dish along the thin
edge of the agar to reduce crowding.
•Dishes were sealed and placed in a vertical position
under the LED and Fluorescent lights at a distance of
34cm. On a photoperiod of 16 hours light and 8 hours
dark.
•Each seed was individually identified.
•Germination rates and root length were measured
weekly for two weeks.
•Dishes were placed on a scanner and the images were
analyzed with ImageJ software; root lengths were
measured.
Summary & Status of Hypotheses
Example of Petri Dishes
Explanation of Graph
The circles show the average root length
and the error bars show 95% confidence
intervals. Root growth from week 1 to
week 2 was greater for plants grown under
Fluorescent lights than for plants under
LED lights (p=0.002, Chi Square
Fdf=1,36=1.8. )
We did not graph the growth from Week 0
to Week 1 because during that interval
growoth was likely produced by energy
from the seed’s endosperm and not from
the light.
Our experiment showed that fluorescent lights
produced plants with longer roots than the LED
light panel. Of course, one of the drawbacks of
fluorescent lights is that it requires more energy
than LED lights to operate. This may be
contribute to higher operational costs, but may
have aided in the difference in root lengths
between the fluorescent and LED lights. Because
the fluorescent lights used more energy, perhaps
that they emitted more energy. This extra energy
could have given the plants grown under
fluorescent lights the advantage. In comparison,
because the LED lights used less energy, they
emit less energy; this could have contributed to
the shorter root length. Since the LED lights
were less intense, we should have moved them
closer to the plants in order to have the same
intensity as the fluorescent lights. In the future, it
would be best to figure out the distance from the
lights to the plants in order to equalize the
intensity being exposed to the plants.
Acknowledgements
We thank G.R.. Davis for assistance with
experimental design , Ab Abercrombie
for help with statistical analysis, and A. Steadman and
T. Player for helpful comments.
Does Seed Depth of Arabidopsis Affect Germination or the Ability to Breach an Agar Surface?
Morgan Hiler, Carrie Martin, and Hannah Leirmoe
Wofford College, South Carolina
Introduction
Arabidopsis seeds are wind distributed; they germinate
and grow where they land in the surface of soil. In the
laboratory to facilitate the measurement of shoot and root
growth the seeds are placed on agar. As far as we know
there have been no previous experimentation to determine
if the seeds of Arabidopsis grow when planted beneath
the surface of agar.
Germination Results
Discussion
Of the 42 seeds planted beneath the domes of agar 7 failed
to germinate, and of the 42 seeds planted on top of the agar
7 also failed to germinate.
There was no significant difference in the germination
rates in either treatment (P-value= 1, Fisher’s Exact Test).
These results support the original hypothesis that the
seeds planted in both conditions would germinate;
however, those seeds planted beneath the agar domes
generally did not break the surface. Therefore, in the
natural habitat of the Arabidopsis, if the seeds are
planted rather than wind distributed the seeds may not be
able to survive. This is because the seeds may not have
enough stored energy to support the plant long enough
for the shoot to break the surface.
Shoot Emergence Results
Hypothesis
Arabidopsis seeds will germinate as well under the
agar as on top of the agar.
Seed Placement
The shoots of the Arabidopsis seeds planted beneath
the agar will not break the surface.
Week 2 Shoot Growth
Methods
Experiment Procedure:
1. Each of the 14 seeds, Ecotype Columbia
Arabidopsis, spaced evenly across the surface of six
Petri dishes filled with 25mL solidified 0.8% agar.
2. Make 42 domes, approximately ½ in in diameter,
with 0.8% agar on a flat.
3. Allow domes a few minuets to solidify, then place
them on top of seven seeds in each Petri dish.
4. Seal dishes and placed them under florescent lights
for a 16 hrs light and 8 hrs dark cycle.
5. After a week check seeds with a dissecting
microscope for germination.
6. After two weeks check seeds again with the
dissecting microscope for germination, and if any
shoots have breached the surface of the agar.
Of the forty-two seeds that were planted beneath the domes
of agar four breached the surface. The other seeds grew
sideways between the bottom of the agar dome and the top of
the agar in the Petri dish. The shoots were beginning to grow
leaves, but they were not able to grow straight up through the
agar.
The seedlings that emerged from the dome’s surface had
been inadvertently displaced, and therefore had a shorter
distance to grow to breach the surface. The seeds that
were directly beneath the center of the domes did not
reach the surface.
These results suggest that if the experiment had
continued the plants may have eventually broken the
agar surface, or died due to insufficient gas exchange.
Our plants did not die in two weeks because unlike soil,
agar is clear and the seeds may have been able to carry
on photosynthesis whereas in soil they could not. The
agar also allows for the shoots to respond to
phototropism from the fluorescent lights rather than just
geotropism, which is all the seeds could respond to if
planted below the surface of soil.
References
Weigel, D. & J. Glazebrook. Arabidopsis: A Laboratory
Manual. 2002. Cold Spring Laboratory Press.
Acknowledgements
Thanks to Professors Davis and Abercrombie for their
support, suggestions, and help with statistical analysis.
Thanks also to T. Player and A. Steadman for their
helpful comments.
The Effects of Agar Concentration on Root Length
in Arabidopsis thaliana
Matthew Boggs, Aaron Seigler, and James Skinner
Department of Biology, Wofford College, Spartanburg, SC 29303
Indroduction
Agar, a gelatinous substance derived from red algae, is
commonly used at a concentration of 0.8% to grow
Arabidopsis plants in the laboratory. To determine
whether this is the optimal agar concentration for root
growth, we measured root growth in agar that was twice
(1.6%) and half (0.4%) the standard concentration. The
Arabidopsis thaliana ecotype Columbia was used for the
experiment as it’s the most commonly used ecotype in
the lab.
Results: Effect of Agar Concentration on Root Growth
Agar concentration had no statistical effect on root growth at 2 weeks
(p= 0.062, Repeated measures analysis of variance, DF=2, 29 F= 3.06)
Status of Hypotheses
1. Agar composition did not significantly affect
root growth in the Arabidopsis thaliana
(Columbia).
2. The density of the agar solution did not affect
germination rates.
Discussion
Hypotheses
The 1.6% agar produced some of the longest
roots, but statistical analysis showed no
significant difference across the agar
concentrations.
The consistency of the 0.4% created an
unstable substrate, such that measurements were
difficult to obtain.
While there was no significant differnce for
any of the concentrations tested, given that the
“p-value” approaced statistical significance, we
feel that a repeat of the experiment would be
appropriate with a larger number of seeds.
Other possible experiments could include
testing of even denser concentration to determine
if density of the solution inhibits root growth.
Nonetheless, we would recommend that the
densities tested in future experiments should not
be below 0.8% to ensure a stable substrate.
1. The greater the density of an agar solution that
Arabidopsis thaliana (Columbia) seeds are cultivated in,
the longer the roots.
2. The density of the agar solutions will have no effect
on germination rate.
Materials and Methods
Materials
6 round agar plates
48 Arabidopsis seeds ecotype (Columbia)
1 set of fluorescent grow lights
0.4% agar in two plates(Control Group)
0.8% agar in two plates
1.6% agar in two plates
Methods
Eight seeds were evenly spaced on upper edge of the
agar in each plate.
Dishes were sealed and marked.
Dishes were placed vertically 33 cm beneath a lamp on a
16 hr light 8 hr dark cycle.
Plates were scanned (below) and measurements of root
length were made weekly for two weeks with Image J
software.
Note: bars are 95% confidence interval.
Results: Effect of Agar Concentration on Germination
Agar concentration had no effect on germination rate (p=0.262, Pearson Chi
Squared Test df2=5.259, data not shown.)
Acknowledgements
The authors are indebted to G. R. Davis for
assistance with experimental design and Ab
Abercrombie for assistance with statistical
analysis.