MITOSIS

Rahel Yacob | Biomedical Science | 26/11/2018

Introduction
CELL DIVISION
The formation of new identical daughter cells. Cell division is vital for;

i. Reproduction –single celled organisms can only reproduce via cell division.

ii. Growth and development – as organisms grow, numerous cells get damaged or die; hence, cell
division replaces these damaged/died cells. Cell division permits growth and mutation by increasing
the total number of cells in the body.

iii. Tissue renewal – damaged or wounded tissues can be regenerated through cell division. [Dinh, H.,
(2018)]

MITOSIS
The process of cell division is called Mitosis, there are five phases of cell cycle; these can be identified by:

i. Interphase– cell is preparing to undergo cell division, at this stage nothing is visible.

ii. Prophase- chromosomes condenses into sister chromatids containing two identical genetic information
gets composed- making it visible.

iii. Metaphase– nuclear envelope disappears, chromosomes then align in the equator independently creating
metaphase plate.

iv. Anaphase– spindle fibres (made up of microtubules) splits the sister chromatids towards the opposite
poles – making daughter chromosomes.

v. Telophase– spindle fibres disappears as the chromosomes has reached the opposite poles, two nuclear
envelope stars to forms. [Yourgenome.org (2017)]

OBJECT
Garlic root was used as an object to observe. Root tips was specifically used due to rapid growth, hence,
numerous cell division and stages takes place.

AIM
To identify interphase and four mitotic stages under microscope.

(218)
MATERIALS AND METHODS
Materials required –

 Solution of Ethanol/Acetic acid (ratio 3:1)

 Hydrochloric acid (2ml)
 Water bath 60
 Cold water
 Feulgen stain solution (2ml)
 Scalpel blade
 Tweezer
 Containers
 Microscope
 Garlic roots
 Glass slides and cover slips
 Goggles
 Lab coat
 Gloves
Methods used-

For the cell division to be visible under microscope, it is important to stain the object in
this case – the chromosomes.

Fix the root tips in Ethanol/Acetic solution (ratio 3:1); fixing the root means that the
organisms are dead and can no longer undergo any further division. Ethanol/Acetic
solution is a fixture that stabilises the structure of the object whilst avoiding any structural
or chemical changes –this ensures that nothing will change during the staining process.

Then relocate the root tips to a bijoux containing 2ml of Hydrochloric acid and place in
the water bath for approximately 10 minutes at 60 Celsius. This process necessary as
Hydrochloric is used to weaken the texture of the tissue – making it easy to press and
cover during observation.

Once the time is up, transfer the root tip in a bijoux containing 2ml of clear cold water
and give it a wash to get rid of Hydrochloric. Then using another bijoux containing
Feulgen stain of 2ml solution, transfer the root tip; leave it to stay in the solution for 15
minutes – no less or more. If less time is given the cells might be not be stained enough to
clearly observe, if more time is given the cells might be overly stained which then makes it
unclear to observe.

The root tip which was originally white/light grey in colour will be dyed to dark purple as
part of being stained, once the 15 minutes is up place the root tip in glass slide using a
tweezer. Beware not to touch.

Using a scalpel blade, cut just the very tip of the root roughly around 2mm and removed
the rest of the root. The tip of the root is the targeted as it is where numerous cell division
occurs. Once successful, place a cover slip on top of the root tip; hold the glass slide and
cover slip vertically and compress using intense pressure but ensuring there is not side to
side movement.

Place the glass slide in the microscope, and ensuring to start with low magnification i.e.
x10; look at the slide and find the root tip. Once found, change to higher magnifications
x40 and/or x100. Always remember of adjust the focus and light to ensure good visibility of
the mitotic stages and gently move around find all he phases.

RESULTS
DISCUSSION AND CONCLUSION
All 4 mitotic phases and the interphase was indeed observed exactly as described; after
strictly following the methods.
However, the process/method was repeated 3 times in order to get the given result;
multiple errors was made which included but was not limited to – drying out of the
root tip, not using the very tip of the root and not squashing properly. The faults were
constantly being acknowledged after successfully staining the Garlic root –
assumptions aroused of possible errors as no mitotic stage; in fact no cells could be
observed.

The method doesn’t necessary specify the time it takes of the roots to dry out and
whether the cutting of tips should be immediate – this may have been the issue that
led to unsuccessful trial. Following the methods mentioned above, individuals could
take their time cutting the tip which will cause drying of the root.
Other problematic issue could be ‘compress using intense pressure’, some individuals
may compress the roots with such a strong pressure that
Feulgen solution is a solution used to only stain deoxyribonucleic acid (DNA) and/or
chromosomes, in the presences of Feulgen solution the chromosomes are outlined in
dark purple/red colour; it enhances the visibility of chromosomes strucure. This
solution is most used in biology, in genetics especially – usually in histology. This
solution has recently been used to detect haematology i.e. mitotic abnormality.
(GARDIKAS and ISRAELS, 1943)
Methylene blue - stains animal cells to make nuclei more visible
Osmium tetroxide - used in optical microscopy to stain lipids black. [Bruckner, M. (n.d.)]

3. Discuss what the limitations of your results are, and the limitations of the
conclusions based on them – the validity of conclusions is dependent on the
experimental method used and the quality of the results obtained.

4. Comment on any limitations of the method, and suggest what modifications could
reasonably be made to the procedure or the apparatus to improve your findings –
immediate cuuting o the root should added andin case drying what alternatives can
be used. For this specific lab report you may wish to research what structures are
being stained with Feulgen solution and what other stains could be used to stain the
same or other structures.

5. State a conclusion and support it with evidence from the data and your knowledge
of biological principles. Compare your conclusions with current published theories
and what other researchers have found. For this specific lab report you may want to
address the importance of determining the Mitotic index of a tissue after you have
researched the topic through the literature
This section should be 200-300 words long and well written, competently discussing
your own results and placing them in context of the current scientific knowledge
Discussion section should contribute 30% to the overall grade for the lab report
References/Bibliography

Dinh, H., (2018). Three Reasons Why Cell Division Is Important. [online] Sciencing.
Available at: https://sciencing.com/three-reasons-cell-division-important-
8289209.html [Accessed 27 Nov. 2018].
GARDIKAS, C. and ISRAELS, M. (1943). THE FEULGEN REACTION APPLIED TO
CLINICAL. [online] Jcp.bmj.com. Available at:
https://jcp.bmj.com/content/jclinpath/1/4/226.full.pdf [Accessed 3 Dec. 2018].

Bruckner, M. (n.d.). Microscopy. [online] Microscopy. Available at:

https://serc.carleton.edu/microbelife/research_methods/microscopy/index.html [Accessed 3
Dec.2018]

SAPS Associates' community (2018). Acetic alcohol and root-tip mitosis. [online]
Saps.org.uk. Available at: http://www.saps.org.uk/saps-associates/browse-q-and-a/391-
what-is-the-purpose-of-the-acetic-alcohol-farmers-fluid-is-in-the-root-tip-mitosis-
investigation [Accessed 28 Nov. 2018].

Yourgenome.org (2017). What is mitosis?. [online] Yourgenome.org. Available at:

https://www.yourgenome.org/facts/what-is-mitosis [Accessed 29 Nov. 2018].
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