Laboratory 4: Preparing and Observing Mitosis in Onion Root Tip Cells Flashcards
Objective
At the end of this laboratory activity, the students will discover the appearance and organization of plant cells in different phases of the cell cycle. Moreover, to visually examine stages of the cell cycle and identify apoptotic changes in cells, and to analyze data to understand the prevalence of each phase and morphological changes during apoptosis.
Materials
- Microscope
- Slides and coverslips
- Fresh onion root tips
- scalpel or razor blade
- 1M hydrochloric acid (HCl)
- Aceto-orcein or methylene blue stain (or similar)
- Beaker
- Warch glass
- Glass pipette or dropper
- distilled water
What are the visible differences between interphase and the stages of mitosis (prophase, metaphase, anaphase, telophase) under the microscope?
- When observing onion root cells under the microscope, it can be seen that during interphase, the nucleus remains intact, with diffuse chromatin and a visible nucleolus.
- In prophase, chromatin condenses into distinct chromosomes, and the nucleolus disappears.
- By metaphase, the chromosomes align at the center, with spindle fibers extending to opposite poles.
- During anaphase, the sister chromatids separate and move to opposite poles, appearing V-shaped as the cell elongates.
- In telophase, the chromosomes decondense, the nuclear envelope reforms, and a cell plate becomes visible, dividing the cell into two daughter cells.
How can you distinguish between prometaphase and metaphase in your sample?
In prometaphase, the nuclear envelope disappears, chromosomes condense but remain scattered, and spindle fibers attach to their kinetochores. In metaphase, chromosomes align at the metaphase plate, with spindle fibers fully developed and symmetrically extending from opposite poles.
Why are onion root tips ideal for studying mitosis? What region of the root contains actively dividing cells?
Onion root tips are ideal for studying mitosis because these roots are regions of rapid growth. As a result, there are many cells actively going through the stages of mitosis, making it easier to observe and identify each stage under a microscope. Additionally, the cells are relatively large and their chromosomes stain very well, allowing clear visibility during microscopic examination (Ray, 2014). The region of the root that contains the actively dividing cells is called the meristematic region. This region is located near the root tip and is responsible for the growth of the root. Cells in the meristematic region are constantly undergoing mitosis to facilitate root elongation (Chaffey, 2014).
Based on your observations, which stage of mitosis is most frequently observed? Why do you think this stage is more common?
Based on the observations, interphase is the most frequently observed stage, followed by prophase. This is because cells spend the majority of their time in interphase, where they grow, replicate DNA, and prepare for division (Chaffey, 2014). Even in the actively dividing onion root tip, most cells are in interphase to ensure they are adequately prepared for mitosis. Prophase is the next most common stage, as it marks the beginning of mitosis and is relatively long compared to the subsequent stages, allowing more cells to be captured in this phase during observation.
What does the proportion of cells in interphase versus mitosis suggest about the duration of these stages in the cell cycle?
The proportion of cells in interphase versus mitosis suggests that interphase occupies a significantly longer duration in the cell cycle compared to the mitotic phases. Since interphase is where the cell grows, replicates its DNA, and prepares for division, it encompasses several subphases (G1, S, and G2) that collectively take up the majority of the cell cycle. In contrast, the stages of mitosis (prophase, metaphase, anaphase, and telophase) are relatively short, occurring quickly to ensure efficient cell division. This aligns with the study of Matagne (1968) and Bryant (1969) about the duration of the mitotic cycle in onion cells. The higher proportion of cells in interphase reflects the extensive preparation and maintenance activities required for a cell to successfully undergo mitosis and indicates that mitosis is a brief event compared to the overall cell cycle.
Why is hydrochloric acid used in the slide preparation process? How does it affect the cells?
Treating the root tips with hydrochloric acid (HCl) helps soften cell walls by partially hydrolyzing cellulose, enabling easier squashing of cells for microscopic observation. It also denatures proteins and breaks down the middle lamella, separating individual cells. Additionally, HCl enhances stain absorption by increasing cell permeability, improving the visibility of chromosomes and mitotic stages. This ensures clear and effective cytological analysis.
Why is the root tip squashed after staining, and what precautions must you take while doing this?
The root tip is gently pressed on the eraser end of a pencil to flatten out the cells in an even layer for microscopic view without overlapping. This avoids the overlapping and makes the chromosomes easy to see. Pressure should be lightly applied, and the cells must be placed between two slides with a coverslip so the pressure can be evenly distributed so that air bubbles are not trapped under the coverslip.
What might happen to the cell cycle if errors occur during mitosis? How could this affect an organism’s growth or health?
Errors during mitosis, such as improper chromosome segregation, can result in aneuploidy (uneven chromosome numbers) or polyploidy (extra chromosome sets). These abnormalities disrupt genome stability and proteome balance, leading to alterations in cellular processes like proliferation. In humans, aneuploidy is associated with conditions such as cancer and developmental disorders, while polyploidy in plants may enhance evolution and crop yields. The cellular response to these errors varies based on tissue type and context, highlighting the complexity of their impacts on growth and health (Storchova, 2021).
How does observing mitosis in onion root tips help us understand cell division in other eukaryotic organisms, including humans?
Observing mitosis in the onion root tip would give insights into cell division- a basic and conserved process in all eukaryotic organisms. In the onion root tip, cells are dividing in an area of active growth, making it the ideal model for studying all phases of mitosis under a microscope. The mechanisms involved in chromosome condensation, spindle formation, and chromatid separation occurring in onion cells are similar to those in human cells, shared by conserved proteins and processes regulating the cell cycle in them. A comparative study like this would help understand how a fault in the cell division cycle results in humans in conditions such as cancer, offering a ground for the development of therapies to correct those errors.
How might environmental factors (e.g., toxins or radiation) affect the mitotic rate in onion root tips? How could you design an experiment to test this?
Toxins and radiation can affect the mitotic rate in onion root tips in an enormous way. For instance, some toxins may cause DNA damage and result in delays in the cell cycle and low mitotic rates. Other toxins might induce the cell to grow out of control, which then causes abnormal growth (Sheddy et. al, 2023). To test these effects, an experiment may expose onion root tips to several concentrations of a given toxin or radiation and compare the mitotic rates of the experimental group to the control group. This helps to understand the effects of environmental stressors on cell division.
How does cytokinesis differ in plant cells (like onions) compared to animal cells? What structural differences explain these variations?
Cytokinesis, the final step of cell division, differs significantly between plant and animal cells due to structural variations. In plant cells, a rigid cell wall surrounds the plasma membrane, which necessitates a unique mechanism for cytokinesis. Instead of pinching inwards, as in animal cells, plant cells form a cell plate in the center during cytokinesis. This plate is created from vesicles containing cell wall materials that fuse to form a new wall between the two daughter cells. Eventually, the cell plate matures into a complete cell wall, separating the two daughter cells.
In contrast, animal cells, which lack a cell wall, undergo cytokinesis through the formation of a cleavage furrow. This furrow is initiated by a contractile ring of actin and myosin filaments, which constricts the cell membrane inward, effectively “pinching” the cell into two separate daughter cells. The process is dynamic and relies on the flexible nature of the plasma membrane, allowing it to deform as the furrow deepens.
These structural differences—namely, the presence of a cell wall in plant cells and the flexible membrane in animal cells—account for the distinct methods of cytokinesis in these two cell types. This divergence in process reflects the adaptations of plant and animal cells to their respective environments and structural constraints.
