Observing Mitosis

Core Practical 5 – From Topic 3 (Voice of the Genome)

Aim: To be able to prepare and stain a root tip to observe the stages of mitosis under a microscope.

Equipment:

  • Garlic roots
  • Sharp knife
  • 1moldm¯³ of  hydrochloric acid
  • Acetic alcohol
  • Orcein ethanoic stain
  • Ice-cold distilled water
  • Water bath (60 ̊C)
  • 2 watch glasses
  • Test tube
  • Test tube rack
  • 2 pipettes
  • Microscope slides
  • Forceps
  • Mounted needle
  • Filter paper
  • Microscope with x100 & x400 magnification

 

Method:

  1. Place a test tube of 2cm³ HCl (1moldm¯³) into a test tube rack in a 60 ̊C water bath.
  2. Cut off 1-2cm of a root tip from garlic root. Put the tip in a watch glass containing 2cm³ of acetic alcohol for at least 12 minutes.
  3. Remove the tip and then place into another watch glass containing 5cm³ of ice cold distilled water. Leave for 4-5 mins, and then remove and dry.
  4. Place the tip into the heated HCl for 5 minutes then repeat the process again by placing tips back into acetic alcohol etc. Tip: The tips will be very fragile at this point.
  5. Transfer 1 tip onto a microscope slide and cut 4-5mm from the growing tip (site of mitosis) and keep it.
  6. Gently break up (macerate) the root tip with a mounted needle.
  7. Add 1 small drop of orcein ethanoic stain and leave for 2 minutes.
  8. Add a coverslip and blot with filter paper.
  9. View under a microscope and identify the stages of mitosis.

 

Results & Calculations: The percentage of cells in each stage of mitosis can be observed and noted down. A Mitotic index can also be formed:

Number of cells containing visible chromosomes ÷ Total number of cells in the field of view

Conclusion: Mitosis consists of four different sections: prophase, metaphase, anaphase and telophase.

Prophase – Chromosomes condense (chromatids joined by centromere)
– Spindle fibres join to both centrioles
– Nuclear envelope breaks down

Metaphase – Centromeres attach to spindle fibres at the equator.
– Chromosomes line up

Anaphase – Centromeres split
– One chromatid from each chromosome is pulled to either end of the cell

Telophase – Chromosomes unravel
– Two nuclear envelopes reform

Cytoplasmic division then occurs where the cell surface membrane divides to form two daughter cells. Interphase then takes place (G1, S and G2 sub-phases). The cell cycle then repeats.

Evaluation Points:

  • Low resolution of microscope (systematic error) – use a more detailed microscope with a greater magnification
  • Human error in counting numbers of cells (random error) – take a picture of the image in the lens as this should make counting easier
  • Not enough time in the solutions to enable successful maceration or staining (systematic error) – leave for 5 minutes longer in future trials
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