2-3-5-required-practical-identifying-mitotic-stages

Required practical: Identifying mitotic stages

• Growth in plants occurs in specific regions called meristems

• The root tip meristem can be used to study mitosis

• The root tip meristem can be found just behind the protective root cap

• In the root tip meristem, there is a zone of cell division that contains cells undergoing mitosis

• Pre-prepared slides of root tips can be studied or temporary slides can be prepared using the squash technique (root tips are stained and then gently squashed, spreading the cells out into a thin sheet and allowing individual cells undergoing mitosis to be clearly seen)

Micrograph showing a stained root tip

Apparatus

• Scalpel

• Forceps

• Onion roots

• Gloves

• Ethanoic acid

• Dilute hydrochloric acid

• Water bath

• Spotting tile

• Microscope slides

• Coverslips

• Pipettes

• Acetic orcein stain

• Paper towels

• Optical microscope

Method

• Garlic or onion (Allium cepa) root tips are most commonly used (the bulbs can be encouraged to grow roots by suspending them over water for a week or two)

• Remove the tips of the roots (about 1cm) and place them in ethanoic alcohol

  • This helps to “fix” the tissue and prevent mitosis from continuing in the cells

• Place the root tips in warm (60^oC) dilute hydrochloric acid

  • This separates the cells

• Transfer the root tip to a microscope slide

• Using a pipette, add a suitable stain (eg. acetic orcein, which stains chromosomes a deep purple)

• Place a coverslip on top of the root tip and press down firmly using a paper towel

  • This spreads out the cells so they are only one layer thick

• View the cells under a microscope

Analysis

• Cells undergoing mitosis (similar to those in the images below) can be seen and drawn

• Annotations can then be added to these drawings to show the different stages of mitosis

Limitations

• The preparation of tissue for microscope slides can damage cells and alter their appearance

  • During preparation, a sample is often squashed or stained, which can generate artefacts

  • The occurrence of artefacts can be decreased by more careful preparation of samples

• The size of cells or structures of tissues may appear inconsistent in different specimen slides

  • Cell structures are 3D and the different tissue samples will have been cut at different planes resulting in inconsistencies when viewed on a 2D slide

• Optical microscopes do not have the same magnification power as other types of microscopes and so there are some structures that can not be seen