Mitosis

• G1 phase involves growth and preparation for DNA replication.
• Interphase consists of three stages: G1, S, and G2.
• The cell cycle is the series of events that occur between one mitotic division and another.
• S phase is the synthesis phase where DNA replicates, resulting in two identical copies of chromosomes.
• Metaphase occurs when chromatids line up at the equatorial plate.
• Anaphase begins with separation of sister chromatids into daughter cells.
• Mitosis is the process by which cells divide to produce genetically identical daughter cells.
• Cytokinesis is the final stage of cell division where the cytoplasm divides into two new cells.
• Prophase is the first stage of mitosis where the nuclear envelope breaks down, spindle fibers form, and chromatin condenses into visible chromosomes.
• Anaphase is characterized by the separation of sister chromatids into opposite poles.
• Telophase marks the end of mitosis with nuclear envelopes forming around new nuclei.
• Cytokinesis divides cytoplasm to form two daughter cells.
• G1 Phase - Cell growth and preparation for DNA replication
• S Phase - Synthesis of new DNA molecules from existing ones
• G2 Phase - Preparation for mitosis (cellular division)
• Why are root tips used in this investigation?

  • in plants, cell division by mitosis is restricted to specific tissues, called meristems
  • found at the growing root and shoot tips
  • this practical involves the preparation and observation of dividing cells from the meristems of actively growing garlic root tips
• Why are several roots used?

  • the roots become very delicate during the preparation stage and may decompose
  • using several roots increases the likelihood that one root tip will survive to give a good root tip squash
• Why are the root tips placed in IM hydrochloric acid at 40°C for 5 minutes?

  • the cellulose cell walls of one plant cell are ‘cemented’ to those of adjacent cells by the middle lamella, that contains pectin
  • the acid hydrolyses the middle lamella, allowing the stain to diffuse into the cells and enabling the tissue to be squashed into a layer one cell thick
  • heating the acid to 40°C increases the rate of hydrolysis of the middle lamella
• Why are the root tips placed in toluidine blue stain for 2 minutes?

  • the toluidine blue stain binds to chromatin DNA, staining it deep blue
  • makes the chromosomes visible
  • staining for two minutes allows the stained chromosomes to be clearly defined
• Why are the root tips gently squashed?

  • to produce a layer that is one cell thick
  • prevents one layer of cells from lying on top of another, obscuring the view of the chromosomes
  • the root tip squash is performed gently so that the cells are not ruptured and to prevent the microscope slide and coverslip from cracking
• How could you increase the reliability of estimates of cell size?

  • measure several cells at the same stage in the same field of view and calculate the mean size for that stage
  • use the same parameter when measuring
  • e.g. top to bottom of a cell
• How could you modify the protocol to improve the definition of the chromosomes within the cells?

  • increasing the time spent in HCl
  • increasing the temperature at which the root tips are incubated in the HCl
  • increasing the time spent in stain
  • increasing the temperature at which staining takes place
• What are possible causes of similarities in mitotic indices?

  • Same species of garlic
  • Cloves from the same bulb
  • Grown for the same amount of time under the same specifically named conditions (e.g. light regime, intensity, temperature) prior to the experiment
• What are possible causes of differences in mitotic indices?

  • Environmental gradient in the place where the cloves were grown
  • Grown for different lengths of time
  • Quality of root tip squash leading to difficulty in observing chromosomes in cells
  • Health of the garlic clove tissue affecting root growth
  • Cloves from several different bulbs which may come from a variety of different species
• How might a mitotic index be used in the study and treatment of cancer?

  It could be used to provide information about how much cell division is taking place within a tumour and therefore how rapidly it is growing.
• As your slide is a snap-shot of the cell division occurring in the root tip meristem at one moment, what do the percentages that you calculated suggest about the length of time that the root tip meristem cells spend in each stage of the cell cycle?

  • there will be many cells observed in the stages of the cell cycle that take a long time, giving high percentages
  • the root tip squashes will contain few cells in the stages of the cell cycle that occur rapidly, giving low percentages.
• HCl is used in root tip mitosis practicals in order to:

  • break down links between cells/cell walls
  • to separate cell walls
  • to hydrolyse cellulose/cell walls

  Why?

  • allows stain to diffuse into cells
  • allows cells to be more easily squashed
  • to stop mitosis
• Cells are gently squashed in root tip mitosis practicals in order to:

  • create a single layer of cells
  • to spread cells out

  Why?

  • to allow light to pass through
• How do you find the mitotic index?

  [the number of cells in mitosis] ÷ [the number of cells not in mitosis]
• Why might other students produce different root tip mitotic indices?

  • roots are a different age
  • garlic was grown in different conditions
  • root tips from different plants/bulbs/species
  • different parts of the root tips in their field of view
  • root tips undergo mitosis at different rates
• Environmental variables affecting the growth rate of root tips:

  • increased concentration of glucose (increased respiration)
  • increased concentration of oxygen (increased respiration)
  • increased temperature (increased enzyme activity)
  • increased concentration of phosphate (increased DNA/RNA/ATP)
  • increased concentration of nucleotides (increased DNA synthesis)
• Nondisjunction means that a pair of homologous chromosomes has failed to separate or segregate at anaphase so that both chromosomes of the pair pass to the same daughter cell. This probably occurs most commonly in meiosis, but it may occur in mitosis to produce a mosaic individual.
• An example of mitosis is the clonal expansion of B cells in the immune system. This is important in making plasma cells, memory cells, and therefore antibodies.
• An agent that causes cancer is called a carcinogen. Give three examples of carcinogens.

  Tobacco, alcohol, UV rays
• HOMOLOGOUS CHROMOSOMES:

  Chromosomes are in matching pairs in most cells; one member of each pair comes from the mother and one from the father (in humans there are 23 pairs).
• SEX CHROMOSOMES:

  One pair of the 23 human chromosomes are the sex chromosomes; they determine the sex of the individual. Females have two X chromosomes and males have an X and Y chromosome.
• AUTOSOMES:

  All the other chromosomes apart from the sex chromosomes.
• What happens to cancerous cells?

  They divide repeatedly, out of control. This forms an irregular mass of cells (a tumour).
• What causes cancerous cells to divide uncontrollably?

  Gene mutation / formation of oncogenes.
• What is the difference between benign and malignant tumours?

  - Benign tumours do not spread from their site of origin. - Malignant tumours spread and invade other tissues.