Report 4: Mitosis

INTRODUCTION:

Mitosis is the process that facilitates the equal partitioning of replicated chromosomes into two identical groups. Before partitioning can occur, the chromosomes must become aligned so that the separation process can occur in an orderly fashion. The alignment of replicated chromosomes and their separation into two groups is a process that can be observed in virtually all eukaryotic cells.

Both the alignment and separation processes are the consequence of the chromosomes interacting with filamentous structures, known as microtubules. The microtubules become organized into a biconical array known as a spindle, which forms early in mitosis, and then disassembles as mitosis nears completion. Mitotic spindles are visible in living cells with the polarizing light microscope. Some of the spindle microtubules become attached to the chromosomes at sites known as kinetochores. The kinetochores cannot be seen with the light microscope, but they reside near the place on the chromosome known as its Centromere, which can be observed with the light microscope. There are two kinetochores on each replicated chromosome (one on each chromatid), and when the replicated chromosome splits apart at its Centromere at the onset of anaphase, each daughter chromosome possesses one Centromere and one kinetochore. The linkages between kinetochores and microtubules are thought to be central in controlling both the positioning of the replicated chromosome at the central portion of the spindle during the alignment phase, and in moving the daughter chromosomes apart after they split at their centromeres. The separation of daughter cells from each other is a process known as cytokinesis, and is separate from mitosis. In cytokinesis, animal and plant cells differ considerably from each other. These differences are the consequence of having or not having a cell wall. Cytokinesis in fungi reveals.

LEARNING OUTCOMES:

            After this experiment, we are able to outline the procedure for preparing acetocarmine squash of a root tip to demonstrate the process of mitosis and able to distinguish and examine;

1.    Every stage of mitosis in plant cell including :

a.    Prophase

b.    Metaphase

c.    Anaphase

d.    Telophase

2.    The process and changes that occurred in every stage of mitosis.

MATERIALS:

1.    1 molar (M) hydrochloric acid (HCL) in dropping bottle

2.    70% ethyl alcohol

3.    Absolute ethyl alcohol

4.    Acetocarmine stain in dropping bottle (dissolve 1.0g in 100mL of 45% acetic acid, boil 5 min, cool, decant and filter)

5.    Alcohol lamp or slide warmer

6.    Beaker in which to let onion sprout

7.    Cover slip

8.    Dissecting needles

9.    Forceps

10. Glass rod for crushing root tips

11. Microscope

12. Scalpel or razor blade

13. Slides

14. Supply of onion bulbs

15. Watch glass

METHOD:

Mitosis in onion root tips

1.    Firstly, we soften the root so that the cells can be separated and flattened, thus making it possible to see the chromosome, nuclei, spindle and other cell parts.

2.    Several drops of 1 molar HCL was placed in a watch glass safely because don’t want the acid get to skin and cloth.

3.    The terminal 3 or 4 mm of the 1 cm long onion root was placed into the acid.

4.    The root tips will feel soft when touched with dissecting needles in short times.

5.    The softened root tips was pick up using the forceps and transferred it to a drop of acetocarmine stain on a clean slide and wait about 15-30 minutes.

6.    The root tip most 1mm was cut and retains by using the razor blade or scalpel. The rest of root was discarded. The remaining root tips were chopped into many pieces with the razor blade and crush the material with glass rod.

7.    A clean cover was applying to the slide and gently heated over the Bunsen burner.

8.    The slide was examined under the low power and high power.

Mitosis in prepared slide

1.    By using the prepared slide, we compared the slide that we did and the prepared slide.

Question and answers:

1.    Most of the cells are in what stages?

In interphase stage.

2.    Can you determine the number of chromosome in any cells?

No

3.    Can you see the spindle in any cells?

Yes

4.    At late telophase, can you see the cell plate?

Yes

5.    Do you observe more cells in mitotic stages than you observed in the untreated root tips?

yes

6.    How does the prepared slide differ from ones you prepare?

It not much differ but slightly differ because do not crashed the onion into many pieces

7.    What advantage do your slides have over the commercial ones?

Slide is fresh prepared compared to the commercial ones might have some damages.

8.    What are the advantages of the commercial slides?

Can use many time and last long period.

9.    What is acetocarmine? What is the function of this solution?

Acetocarmine is a staining that colors the chromosome to make the chromosome more visible through microscope.

DISCUSSIONS:

            Mitotic division consists of two, meiosis and mitosis. The onion root tips occurs the mitosis process. In mitosis process there are 5 stages, Interphase, Prophase, Metaphase, Anaphase and Telophase. Why do we observe the root tips of the onions? This is because in the root tips there were actively divided. The onions bulbs supported by tripod of toothpicks in beaker of water.

            Most of the cells are spent in the interphase stages. Interphase consist of G₁ (growth phase or gap), S (synthesis phase) and G₂ (second growth phase or gap 2) then followed by mitosis. In G₁  volume of cytoplasm increase and the protein start to synthesis. Increase the number of organelles such as mitochondria, endoplasmic reticulum and Golgi apparatus. Also synthesis of carbohydrate lipid and ribonucleic acid (mRNA, rRNA and tRNA). S phase is DNA synthesis phase. The cell DNA replicate and chromosome duplicate. In this stage in which genetic materials are synthesize. In G₂ the energy stored are increase. The formation of microtubules (kinetochore, aster and polar. Increase in cell organelles (mitochondrion, Golgi body and endoplasmic reticulum) also increase in size of nucleus. 

            Mitosis begins with Prophase. In Prophase, the chromatin fibers more tightly coiled, condensing into discrete chromosome. In the same time, the nucleoli were disappeared. Each duplicated chromosome appears as two identical sister chromatids joined together. The mitotic spindles begin to form. It is composed of the centrosomes and microtubules that extend from them to form spindle. The radial arrays of shorter microtubules that extend from the centrosomes are called asters. The centrosomes move away from each other, apparently propelled by the lengthening microtubules between them.

            In metaphase, the centrosomes are now at opposite ends of cell. The centrosomes convene on the metaphase plate. The chromosomes, centromeres lie on the metaphase plate. For each chromosome, the kinetochore of the sister chromatids are attached to the kinetochore microtubules coming from opposite poles. The entire apparatus of microtubules is called the spindle.

            In anaphase, it begin when 2 sister chromatids of each pair suddenly apart. When the microtubules shorten, the centromeres splits and sister chromatids are pulled to different poles. By the end of anaphase, the 2 ends of the cell have equivalent and complete collection of chromosomes.

            In Telophase, nucleolus is forming. 2 daughter nuclei begin to form in the cell. Nuclear envelope arises from the fragment of the parent cell nuclear envelope and the chromosome less condense.

            Cytokinesis is the division of the cytoplasm is usually well underway by late telophase, so the daughter cell appears shortly after the ends of mitosis. In the onion root tips cells, it has the cell wall, so during the telophase, the vesicle derived from the Golgi apparatus which contain carbo-filled vesicle move along the microtubules to the middle of the cells producing the cell plate. The cell plates enlarge until its surrounding membrane fuses with the plasma membrane along perimeter of the cell. Two daughter cell result, each with its own plasma membrane. A new cell wall arising from the contents of the plate has performed between the daughter cells.

The mitotic index:

Interphase: 

Prophase:

Metaphase:

Anaphase

Telophase

Conclusion:

            As a conclusion in the root tips cell that I observed are all active divided because of the mitotic percentage of each cells are more than 0.05%. We also able to distinguish each stage in mitosis process (interphase, prophase, metaphase, anaphase, telophase). We also know the function of the acetocarmine and the function of HCL.

REFERENCES:

Brooker, R. J. (2009). Genetics Analysis & Principle. Third edition: Mc Graw Hill international edition.

Eldra P. Solomon, Linda R. Berg, Diana W.Martin. (2005). Biology. Third Edition: Thomson Brooks/Cole.

Kenneth R. Miller, joseph Levine. (2004). Prentice Hall Biology. Teacher's Edition: Pearson Publishing.

Neil, C. O. (2005). Biology. Seventh Edition: Benjamin Cumming.