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Research Question:
“The important thing is not to stop questioning. Curiosity has its own reason
(Calaprice, 2000)
Abstract
plants, such as space, are limited. Hence, the optimisation of plant growth is
importunate to ensuring that our reserves are used efficiently. The onion is the
but historic role in diets and traditional medicine. NPK fertilisers are frequently
used to augment plant growth, but their ratios are varied, each theorised to
affect plants in different ways, with limited supporting evidence. Hence, the
2:1:1, 2:2:1 and water) in aqueous fertilisers affect the growth of the
20 onions bulbs were placed in 4 hydroponics set ups, each holding 5 bulbs
and a different solution. This inquiry was explored through the recording of the
mitotic indices (%) of onion root tips, taken from bulbs placed in different
solutions, over a period of 5 weeks. 1:1:1, 2:1:1 and 2:1:1 NPK solutions were
tested due to their prevalent use worldwide and water was used as a control.
The results suggest that a 1:1:1 NPK ratio is optimal for onion growth (the
paired t-tests indicated significant differences between the 1:1:1 solution and
remaining NPK solutions). The water solution resulted in a high mitotic index
similar to that of the 1:1:1 solution, but due the propensity of the water
Table of Contents
Abstract_______________________________________________________i
List of Abbreviations_____________________________________________v
List of Figures_________________________________________________vi
Chapter 1: Introduction
Chapter 2: Investigation
2.1: Hypothesis………………………………………………….…….5
Chapter 3: Method
3.2:Organisms…………………………………………………...…..10
3.4: Limitations……………………………….…………………...….12
iv
Chapter 4: Results
Chapter 5: Discussion
5.3: Conclusion……………………………….……….…………….24
References…………………………………………………….…………….27
Appendices
List of Abbreviations
List of Figures
24/11/2015…………………………………………………………………………11
micrograph………………………………………………………...…………….…11
Figure 3: A bar graph displaying how the average mean mitotic index (%) of
all replicates varied between the 4 solutions tested, with each series referring
to a replicate……………………………………………………..…………………16
Figure 4: A bar graph displaying how the average mean mitotic index (%)
SD.………………………………………………………………..…………………20
1
1: Introduction
For millennia, plants have been part of the human diet as food or
The onion (Allium cepa) is commonly utilised for such dietary and
The Tamils in India have a similar belief (Grubben, 2004). Onions are rich in
agents) (NOA, 2011). These benefits may have been the source of the
mentioned beliefs. The consequent high demand for onions would be better
met by a high rate of onion growth. Hence, I started looking into how onion
made of specialised proteins that facilitate ATP production (Ng, 2013). For
producing ATP, which releases energy when hydrolysed (Ng, 2013). The
fertilisers. This also creates a low turgour pressure that facilitates mitosis
(Sano, 2007).
important role in the cell cycle. This is substantiated by the fact that a
different NPK ratios on the growth of onions. In fact, NPK fertilisers are often
used to supplement plant growth. However, they vary in their elemental ratios
(N:P:K) and effects of different ratios on growth have not been fully explored,
How do different NPK ratios (1:1:1, 2:1:1, 2:2:1 and water) in aqueous
fertilisers affect the onion growth (Allium cepa) over a 5-week period?
Multiple studies have shown that using NPK fertilisers can result in a
rostrata (Becker, Diekmann, 1991). This indicates that the nitrogen stimulated
the root nodules, increasing the rate of nitrogen fixation, hence increasing the
fertiliser has also reportedly been linked to increased okra (a Nigerian crop)
study proved that crops planted in nitrogen deficient mediums had decreased
impeded growth.
(1985) and more reliable as it considers modern concepts that had replaced
older paradigms.
onions, using hydroponics. I hope that this investigation will promote further
results.
production of onions per unit time and bring greater economic benefits.
2: Investigation
2.1: Hypothesis
I hypothesize that the 2:2:1 and 2:1:1 ratios will produce the highest
mean mitotic indices across all replicates, because these 2 ratios have the
highest nitrogen contents of all the tested ratios. A higher quantity of available
nitrogen would likely promote greater growth per unit time. As nitrate ions
polypeptides that facilitate growth, like the hydrolase enzyme invertase, which
onion bulbs rather than onion seeds would be more suitable for the
experiment. The method employed is identical to that in section 3.3, but only 2
pots were used, one with 5 onion bulbs and one with 5 seeds. Both pots
6
supported 1 seed while 4 sponges supported one bulb. The final method used
was adjusted in accordance with the findings from this preliminary experiment.
difficulty in determining the mitotic index of the seeds, as few cells had
the cell cycle that each cell was in could not be distinguished. Secondly, more
sponges cubes were used to support the bulbs as it was observed that the
The independent variable was the NPK ratio in each solution. The NPK ratios
tested were (when simplified), 1:1:1, 2:1:1 and 2:2:1. The 1:1:1 ratio was used
due to its reputation as being the best ratio for “all-purpose,” use (Smith,
2013). The higher nitrogen level ratios (2:1:1 and 2:2:1) were chosen because
these ratios are the most common compound fertiliser ratios in the EU
The dependent variable was the mitotic index of the tip of the longest
root of each onion. I chose the root as the tissue to be examined because
enhance overall onion growth, high root growth would indicate high overall
onion growth. This is because larger roots would produce more cytokinin per
unit time, hence facilitating increased growth. Therefore, root mitotic indices
The species of onion used in the experiment was Allium cepa. Different
onion species would respond differently to the same NPK ratio, limiting the
extent to which the different ratios can be compared with respect to the
nutrient uptake by the onions, hence causing different effects on the growth of
placed before the start of the experiment was identical for all the onions as
The time at which the micrograph for each sample was carried out was
kept constant (3:00pm each Tuesday); this was to ensure that the gaps
The buoyancy of all sponges used was the same at the start of the
experiment. By drying the sponges at the start of the experiment, their initial
water contents were identical. Hence, their buoyancies were near identical;
8
therefore, the volume of solution that the roots were initially exposed to was
equal for all the onions as they were held by sponges with near equal water
saturation.
The pots used for each set up were identical in dimensions. This
ensured that the volume of each pot was identical, so that the roots had the
same growth space. If this were varied, the roots would have grown
differently.
9
3: Procedure
Apparatus Uncertainty
4 pots
Tap water
Metal spatula
Metal scalpel
100cm3 beaker
Glass rod
Light microscope
40 glass slides
55.863g of NH4H2PO4
28.878g of KCl
5.232 g of NH4NO3
3.2: Organisms
20 onion bulbs were obtained from the Marine Parade wet market.
Firstly, 4 shallow pots with heights of 4cm and width of 41cm were
(±0.5cm3). Secondly, for each NPK ratio tested, the appropriate masses of
100cm3 beaker was then placed on an electronic mass balance that was set
to zero after the beaker had been placed on it. Using a spatula, sufficient
quantities of each compound were added until the desired mass of each
compound was reached for a particular ratio. This was repeated for all 3 ratios
under investigation. Next, the 4 pots were labelled as “1:1:1,” “2:1:1,” “2:2:1,”
and “Water,” for identification. The corresponding solid fertiliser mixtures were
added to their respective pots. Once added, the mixture was stirred with a
Green polystyrene mesh, held taut by duct tape, was used to cover the
surface of the pots following the addition of the fertilisers. To secure the
onions in place, each onion was placed in 6 fused sponge cubes, each of
length 1cm, with a circular hole cut in the centre using a metal scalpel to
accommodate the base of the onion bulb. 5 onions were then placed on the
mesh covering each pot, to allow for 5 repeats per solution and each were
The tip of the longest root was cut from each onion using the metal
scalpel before the onions were placed into the sponge base. Each root tip
11
distinguished. The samples were placed between 2 glass slides each, and
This sampling and analysis was repeated every week over a 5-week
period to determine the mitotic index of the 20 onion root tips across all
solutions. 250cm3 of water was added to each pot using the measuring
fertilisers.
Figure 2: An onion cell micrograph – Baker, The Stages of the Cell Cycle (Mitosis – Interphase and
Prophase), 2013
12
The stage of mitosis each cell was in was determined by studying their
extensions from centrioles) push the chromatid pairs to the equator of the cell.
either lateral side of the cell, having been pulled by the mitotic spindles. On
either side of the cell, during telophase, nuclear membranes develop around
the chromatids that have started to uncoil. To ensure the investigation was
stage (Allott, 2014) and practised identifying which stage of mitosis a cell was
3.4: Limitations
grown using hydroponics. Its results do not reflect the effect on onion grown in
4: Results
1. 2 onions in the water solution and 1 onion in the 1:1:1 NPK solution
2. The roots in the water solution had become flaccid after 4 weeks.
13
3. Large root networks formed in the water and 1:1:1 NPK solutions, the
4. A white powder was observed on all the pots on the third week –
Data processing was conducted using the raw data in appendix 1.2.
Table 1: A table showing the equations utilised in data processing, with sample
calculations
!"#$%#&'(#) !" !"##$ !"#$%&'("& !"#$%"% Mitotic index of replicate 4 in the water solution
𝑀𝑖𝑡𝑜𝑡𝑖𝑐 𝐼𝑛𝑑𝑒𝑥 = ⋅ 100%
!"#$% !"#$%& !" !"##$
!!!!!!!
⋅ 100%= 6.67%
(Allott, 2014) !"
telophase.
𝑡ℎ𝑒 𝑚𝑖𝑡𝑜𝑡𝑖𝑐 𝑖𝑛𝑑𝑒𝑥 𝑜𝑛 𝑒𝑣𝑒𝑟𝑦 𝑤𝑒𝑒𝑘 (𝑓𝑜𝑟 𝑜𝑛𝑒 𝑟𝑒𝑝𝑙𝑖𝑐𝑎𝑡𝑒) Mean for the replicate 1 in the water solution
𝑀𝑒𝑎𝑛 =
5
= 50.67%
𝑋−𝑋 !
𝑆𝑡𝑎𝑛𝑑𝑎𝑟𝑑 𝐷𝑒𝑣𝑖𝑎𝑡𝑖𝑜𝑛 (𝑆𝐷) = SD for replicate 1 in the water solution
(𝑛 − 1)
(5 − 1)
index, 𝑛 = number of mitotic indices in the row,
= 42.32 (rounded to 2 decimal places)
represents the sum across all values in the row
14
Table 2: A table showing the mitotic index (%) of each onion root tip immersed
in the water solution over 5 weeks including the mean mitotic index for each
appendix 1.1.
Table 3: A table showing the mitotic index (%) of each onion root tip immersed
in the 1:1:1 NPK solution over 5 weeks including the mean mitotic index for
each repeat and standard deviation, using data from tables 2, 6, 10, 14 and 18
in appendix 1.1.
Table 4: A table showing the mitotic index (%) of each onion root tip immersed
in the 2:1:1 NPK solution over 5 weeks including the mean mitotic index for
each repeat and standard deviation, using data from tables 3, 7, 11, 15 and 19
in appendix 1.1.
Table 5: A table showing the mitotic index (%) of each onion root tip immersed
in the 2:2:1 NPK solution over 5 weeks including the mean mitotic index for
each repeat and standard deviation, using data from tables 4, 8, 12, 16 and 20
in appendix 1.1.
Table 6: A table showing the mean mitotic index (%) of each root tip over 5
weeks for the water, 1:1:1, 2:1:1 and 2:2:1 solutions, including means and
standard deviation
Replicates Solution
Water 1:1:1 2:1:1 2:2:1
Mean mitotic Mean mitotic Mean mitotic Mean mitotic
index (%) index (%) index (%) index (%)
1 50.67 57.33 32.67 24.67
2 51.33 59.33 31.33 32.67
3 30.00 35.33 20.00 28.33
4 50.00 66.67 29.33 25.33
5 60.00 45.33 36.67 34.67
Mean (%) 48.40 52.80 30.00 29.13
SD (%) 11.06 12.41 6.20 4.42
80
70
Mean mitotic index (%)
60
Series1
50
Series2
40
Series3
30 Series4
20 Series5
10
0
Water 1;1;1 2;1;1 2;2;1
Figure 3: A bar graph displaying how the average mean mitotic index (%) of all replicates
varied between the 4 solutions tested, with each series referring to a replicate
onions grown in certain solutions were statistically higher (p ≤ 0.05) than the
were chosen because they test if one set is statistically greater or lower,
17
than of set B
Table 7: A table of descriptive statistics for the water, 1:1:1, 2:1:1 and 2:2:1
solutions
Table 8: A table of continued descriptive statistics for the water, 1:1:1, 2:1:1 and
2:2:1 solutions
Table 10: One-tailed t-test results for the first set of tests
Table 11: One-tailed t-test statistics for the second set of tests
Table 12: One-tailed t-test results for the second set of tests
5: Discussion
The processed data clearly indicates that the solution used in growing
(Figure 4).
The water solution yielded a very high mitotic index (48.40%), the
second highest of all tested solutions. However, the data had high SD
The onions grown in the 1:1:1 solution had the highest mitotic index
mitotic index for a repeat was only 35.33%, which is similar to the maximum
The mitotic index of the root tips placed in the 2:1:1 solution had a low
value of 30.00% and low SD (6.202%), suggesting that these onions had
limited growth due to the fertiliser used. This is synonymous with previous
The 2:2:1 solution also resulted in limited root growth as shown by the
mitotic index between the 2:1:1 and 1:1:1 solution; the 2:2:1 and 1:1:1
solution; the water and 2:1:1 solution and the water and 2:2:1 solution (tables
20
9 and 11). This delineates the fact that the water and 1:1:1 solutions beget
between the mitotic indices of onions placed in the 1:1:1 and water solution,
and those of the 2:1:1 and 2:2:1 solutions. Hence, it can be determined that
both high nitrogen fertilisers decrease onion growth to the same extent, and
additionally, that the 1:1:1 and water solutions analogously increase the
70
M3an mitotic index across all repeats
60
50
over 4 weeks (%)
40
30
20
10
0
Water 1;1;1 2;1;1 2;2;1
Type of solution of used
Figure 4: A bar graph displaying how the average mean mitotic index (%) across all
Limitation Modification
system in the hydroponics set ups, remove waste solution and replace
growth.
2. The lack of use of NH4NO3 in the Other compounds could have been
1:1:1 and 2:2:1 solutions resulted in used such that each solution
nitrate ions.
3. The ages of onions used were The onions could have been
varied, as the onions had been planted at the same time and their
grown at different times, hence their bulbs could have been harvested
ability to grow further was varied as after one month for use in the
Limitation Modification
S-phase of mitosis.
Limitation Modification
meshes in the third week. This area of the pots, temperature, wind
onions.
mitotic index.
Limitation Modification
5.3: Conclusion
should be used as a solution because it induces high onion growth. This has
disproven my initial hypothesis that the 2:2:1 and 2:1:1 solutions facilitate
research on NPK ratios in fertilisers and their effect on growth (Brown, 1980;
The two high nitrogen ratios had the lowest mitotic indices. The
attributed to the negative effects of the high nitrogen content of both fertilisers.
Large amounts of ATP were used to absorb nitrate and ammonium ions in the
total energy a plant consumes.” (Chapin, 1987) This therefore reduced the
mitotic index was significantly higher in the water and 1:1:1 solution compared
to the 2:1:1 and 2:2:1 solution, as the water and 1:1:1 solutions did not have
25
The 1:1:1 solution had the highest mean mitotic index as it contained
the necessary nutrients for a high rate of mitosis in sufficient quantities such
that all the nutrients could be absorbed without depleting significant amounts
of ATP. However the difference in mitotic indices between the water and 1:1:1
solution was statistically insignificant. This shows that tap water is a useful
1:1:1 NPK solution, across five weeks. This is possibly due to the multiple
nutrients present in tap water, such as nitrate and phosphate ions, albeit in
However, in the water solution, rot had formed on the roots in the final two
weeks. This reduced the number of living cells and, consequently, the onions’
mitotic indices in the last two weeks of the experiment. This can be attributed
to the shallow root network that had limited access to air, whereas the root
networks of the onions in the 1:1:1 solution were closer to the surface. Hence,
the roots in the water solution had little exposure to air, hindering their ability
to respire, promoting apoptosis (cell death) in root cells and reducing the
mitotic index.
apparatus used and minor impact of the limitations of the experiment on the
results from the NPK solutions. These results are possibly of high significance
References
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Appendices
compared. NH4H2PO4 has 1 nitrogen atom and 1 phosphorus atom, and KCl
has 1 potassium atom, therefore one mole of both compounds would produce
was only used in the 2:1:1 solution, as its inclusion allows for double the
would double). For the 2:2:1 solution, there must be 2 moles of nitrogen and
phosphorus for every mole of potassium and this can be accomplished if the
The appropriate masses of NH4H2PO4, KCl and NH4NO3 needed for each
appendix 1.1.
34
Table 1: A table displaying the number of moles and mass of each compound
As the ratio used in this experiment is a molar ratio, the number of moles of
each compound needed for each ratio is found. This was done in the manner
listed below.
compound required.
𝑚𝑎𝑠𝑠 (𝑔) = 𝑚𝑜𝑙𝑒𝑠 𝑚𝑜𝑙 ⋅ 𝑅𝑒𝑙𝑎𝑡𝑖𝑣𝑒 𝑓𝑜𝑟𝑚𝑢𝑙𝑎 𝑚𝑎𝑠𝑠 (𝑔𝑚𝑜𝑙 !! ), the number
booklet.
35
needed for the generation of a 1:1:1 NPK ratio was found. Following this,
189.59𝑥 = 30
was solved.
a 30 cell area 1 week after the start of the experiment across the 5
a 30 cell area 1 week after the start of the experiment across the 5
a 30 cell area 1 week after the start of the experiment across the 5
a 30 cell area 1 week after the start of the experiment across the 5
a 30 cell area 2 weeks after the start of the experiment across the 5
Table 10: A table showing the number of cells in each stage in mitosis in
a 30 cell area 2 weeks after the start of the experiment across the 5
Table 11: A table showing the number of cells in each stage in mitosis in
a 30 cell area 2 weeks after the start of the experiment across the 5
Table 12: A table showing the number of cells in each stage in mitosis in
a 30 cell area 2 weeks after the start of the experiment across the 5
Table 13: A table showing the number of cells in each stage in mitosis in
a 30 cell area 3 weeks after the start of the experiment across the 5
Table 14: A table showing the number of cells in each stage in mitosis in
a 30 cell area 3 weeks after the start of the experiment across the 5
Table 15: A table showing the number of cells in each stage in mitosis in
a 30 cell area 3 weeks after the start of the experiment across the 5
Table 16: A table showing the number of cells in each stage in mitosis in
a 30 cell area 3 weeks after the start of the experiment across the 5
Table 17: A table showing the number of cells in each stage in mitosis in
a 30 cell area 4 weeks after the start of the experiment across the 5
Table 18: A table showing the number of cells in each stage in mitosis in
a 30 cell area 4 weeks after the start of the experiment across the 5
Table 19: A table showing the number of cells in each stage in mitosis in
a 30 cell area 4 weeks after the start of the experiment across the 5
Table 20: A table showing the number of cells in each stage in mitosis in
a 30 cell area 4 weeks after the start of the experiment across the 5