Onion Cells Measurement

Introduction

Onion epidermal tissue has been used over time as a model to study cell structure
and plant organelle systems because of its transparency and lack of chloroplasts.
As an onions cell is composed like any other plant cell (cell wall, cell membrane
and organelles), the presence of a cell wall allows for a better and clear view of the
general shape and contents that the cell has (Onion and Cheek Cells (Theory) :
Class 9 : Biology : Amrita Online Lab, s. f.). This tissue has been used for several
investigations, such as looking at the mechanics between cells, organelle
localization and their interaction with each other (Scott et al., 1999).

The common onion or Allium cepa is the most used type to investigate the
interaction and structure of cells. This onion, however, has multiple color varieties,
ranging from white to red. Because red onions are cultivars (selected for their
characteristics) ,(C.D Brickell et al., 1981) some other difference could also be
included in that variety. Considering that one of the characteristics used to study
this onion is the translucent peel that can be observed, there could be a difference
between the two varieties because of the different colors they present.

Objectives: Using a microscope, analyze the variability of cell size between two
varieties of common onion (Allium cepa) as well as observe the structure of the
cells found in the onion’s tissue sample.

Investigation question: Will the size and structure of the cells found in the
common onion (Allium cepa) tissue be different when comparing the white and the
red variety of it under a microscope?

Variables:

i. Independent: Type of A.cepa used in the analysis; piece of tissue

used for the observation.
ii. Dependent: Size and structures found after the observation of
multiple pieces of tissue from the varieties of A.cepa.
iii. Control: Indicator used to visualize the different cells and structures
found in A.cepa (Methylene Blue Stain); level of magnification used to
observe the tissues (4X for this experiment).
 Hypothesis: Considering that the tissues to analyze both originated from
the same species of onion (Allium cepa), there will not be any significant
difference on either the size or the structures found when comparing a
sample from the different color variants of the species.

Methodology:

a. Preparation of microscope slides:

i. Place a drop of water in the center of the slide.
ii. Position an onion layer (thinnest) on liquid.
iii. Place one to two drops of stain (methylene blue)
iv. Wait for 40 seconds and rinse the sample with water using the
pipette.
v. At ta 45° angle, place one side of the cover slip against the
slide contacting the outer edge of the water drop.
vi. Lower the cover slowly, avoiding air bubbles.
vii. Remove excess water with the Pasteur Pipette.
viii. Repeat steps 1-5 to prepare 5 samples.

b. Measurement of the Field of View (FOV):

i. Set the objective magnification at 4X.
ii. Position the ruler under the objective lens and measure the
FOV.
iii. Record the data.

c. Observation of the sample:

i. Set the objective magnification at 4x.
ii. Place sample 1 in the stage and lock it using the clip.
iii. Focus the image and count the number of cells in the FOV.
iv. Record the data.
v. Repeat steps 1-3 with samples 2-4

a. Materials
- Microscope Zeiss model Primo star
- 2 Slides and 2 covers.
- 2 50 mL Beakers.
- 100 mL of water.
- Precision Cutting Knife.
- 2 Tared Watch Glasses.
- 1 Ruler.
- 2 Pasteur Pipettes.
 b. Reagents
- 50 mL of Methylene Blue Stain.
- (x) pieces of onion tissue (red and white).

Results

Table 1. Raw data.

Image size Real size

Image size Real size
(cm) (µm)
(cm) (µm)

1,20 141,18
2,70 117,39

1,50 176,47
1,80 78,26
Image Red
Image White
Onion 1
1,70 200,00 Onion 1
1,30 56,52

0,90 105,88
2,50 108,70

1,40 164,71
1,40 60,87

1,40 164,71
1,60 177,78

1,50 176,47
1,30 144,44
Image Red
0,70 82,35 Image White
Onion 2 0,90 100,00
Onion 2

2,20 258,82
1,00 111,11

1,20 141,18
1,40 155,56

0,90 105,88
1,00 117,65

Image Red
1,20 141,18 Image White
Onion 3 0,90 105,88
Onion 3

0,70 82,35
0,90 105,88
 1,20 141,18
1,40 164,71

0,40 47,06
1,30 152,94

1,00 117,65
1,00 117,65

0,70 82,35
1,50 176,47

Image Red 0,50 58,82 Image White 1,10 129,41

Onion 4
Onion 4

1,10 129,41
1,60 188,24

0,80 94,12
0,70 82,35

Table 2. Processed data.

❑x T value T value
x (µm)  (calculated) (tabulated)

Red Onion 130,5882353 50,0380478

11,18884764
0,579630323 2,10
8,33543104
White Onion 122,5902245 37,2771809

Graph 1.
Mean of both varietys of Allium cepa
140

120

100

80
Size (µm)

60

40

20

0
Red White
Variety of onion
 c. Calculations:

Discussion:

Looking at graph 1, it can be seen that the average size of the different onion cells
when being compared with each other don’t have a very high difference in
measurement. Although the average difference is of ~8 µm, after doing the t-
student value analysis, where the result was ~0.58, when compared to the
tabulated value in the statistical analysis using 2 tails and a trust interval of 0.05%
which resulted in 2.10, it can be said that because the calculated value was lower,
the difference between both measurement is not significant enough to state that
the size of the cells is correlated with the variant of onion used in the observations
of this experiment.

Because the strandart deviation from each measurement was different and the one
from the red onion was higher, after doing the statistic error calculus, it could be
stated that there was a higher error when measuring the cell size of this group of
onions. Although the difference was not too much, it was enough to state that
some of the methodology could have been better when observing and preparing
those samples.

As the onions used in the experiment belong to the same species, Allium cepa, the
possibility of the structures of the onion being different is very small. However,
because this onion has been selected to cultivate for its characteristics, if it keeps
being selected for example because of overall size or even hybridized with other
type of onion, some structures could change and the cell size could be one of
them.

Conclusion and Evaluation

The investigation hypothesized that the size of the cells was not correlated with the
variety of the onion used. Because of the results found, the null hypothesis was
rejected by the lack of correlation between the coloration of the onion and a
significant size difference between the cells of the onions.

Taking into account that the methodology for this observation could be more
precise, a bad manipulation of the tissue as in pressing it too hard could to an
extent widen the cell affecting the perception of real size that it would have intact.
On the other hand, depending on the thickness of the tissue used the visibility of
the cells can also be compromised, leading to uncertain measures and possible
error in analysis.

As this is a standard method when observing onion cells, it a very easy and friendly
method to use, error mainly come from user manipulation and lack of attention.
The use of coloration using Methylene Blue Stain allows for a clear visualization of
the cell wall and therefore makes it easier to measure it.

References:

C.D Brickell et al. (1981). INTERNATIONAL CODE OF NOMENCLATURE FOR

CULTIVATED PLANTS 1980. International Association for Plant Taxonomy,

30(1). https://doi.org/10.1002/j.1996-8175.1981.tb00756.x

Onion and Cheek Cells (Theory): Class 9: Biology: Amrita Online Lab. (s. f.).

Recuperado 28 de mayo de 2021, de http://amrita.olabs.edu.in/?

sub=79&brch=15&sim=125&cnt=1

Scott, A., Wyatt, S., Tsou, P.-L., Robertson, D., & Allen, N. S. (1999). Model

System for Plant Cell Biology: GFP Imaging in Living Onion Epidermal Cells.

BioTechniques, 26(6), 1125-1132. https://doi.org/10.2144/99266st04