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Root tip mitosis

A video of a class practical to investigate root tip mitosis
ALEX JENKIN: Unlike most animals, plants have specific zones where growth can occur. Each zone contains a meristem, which consists of an area of dividing undifferentiated cells. Clusters of new cells which are increasing in size, elongation, and slightly older cells which become specialised into different tissues, differentiation. This practical uses the meristems found in the root tips of actively growing garlic roots to observe cells undergoing mitosis.
You will need the following equipment, eye protection, small glass vial, 1 mol of hydrochloric acid, a water bath at 40 degrees C, a Plastic Pasteur pipette, a three-day-old rooted garlic clove suspended over water with a cocktail stick, a cutting tile, a beaker of tap water, 1% toluidine blue stain, dissection scissors, a small watch glass, a microscope slide, cover slips, a mounted needle, paper towels, and a compound light microscope. You can carry out the first part of this protocol using a water bath away from the students so they don’t need to work with the hydrochloric acid. Firstly, half fill a small vial with 1 mol of hydrochloric acid.
Stand the vial in the 40 degree water bath for 15 minutes to allow the acid to reach the temperature of the water bath. Using the cocktail stick, transfer the garlic clove to the vial of acid so the roots are submerged in the acid. Leave the vial containing the garlic clove in acid in the 40 degree water bath for five minutes. The hydrochloric acid kills the cells and breaks down the cell walls and membranes allowing the stain to reach the chromosomes. After five minutes, use the cocktail stick to carefully remove the garlic clove from the acid, and gently rinse the roots in tap water.
Using scissors, cut off the terminal 3 millimetres of the root tips, and allow them to fall onto a small watch glass.
Add one drop of 1% toluidine blue stain. Leave the garlic and the stain at room temperature for two minutes.
Remove the excess stain and rinse the root tips in the watch glass using a plastic pipette and tap water.
Carefully transfer the root tips to a clean microscope slide.
Gently spread the root tips with a mounted needle so that they are not overlapping, and add a drop of tap water.
Carefully, place a cover slip on top.
Wrap a paper towel around the slide and cover slip a couple of times.
And avoiding sideways movement, press down on the top of the cover slip with the thumb to squash the root tips and to spread them out.
View the root tip squash through a light microscope with the times 10, then the times 40 objective. Spend a few minutes looking over the whole of your prepared slide. You’re looking for a single layer of small square cells that are found in rows. They have no obvious vacuole or thickening of the cell walls. You should be able to identify cells that are undergoing different stages of mitosis. The cell cycle is thought to be influenced by the plant circadian clock, so it suggested that active cell division is most likely to be observed in the morning or around midday. This practical allows students to view cells undergoing mitosis using plant tissues that were actively growing moments before the practical commenced.
This is a fascinating and enthralling aspect of this investigation.
Usually when you look at a cell under a microscope, the nucleus just looks like a dark spot and you cannot see the individual chromosomes within it. However, when a cell divides, the chromosomes become visible. When this happens it is possible for students to see how the chromosomes move as the cell passes through each stage of cell division.
Meristems are structures at the ends of roots and shoots where plant stem cells divide by mitosis (cell division) to form identical daughter cells. In this practical, students prepare and observe cells in various stages of mitosis from the meristems of garlic root tips that were actively growing moments before the practical commenced.

Root tip squash

This protocol has been developed in collaboration with CLEAPSS (the school science safety consultants in England) to minimise the risks involved in the stain that is used and in the use of hot acid.
You can grow the garlic roots by using a cocktail stick to suspend the cloves of garlic over water, like this. This also enables the garlic clove to remain intact for the majority of the treatments and minimises the handling of hot hydrochloric acid.
If you want to, you can split the cloves into smaller sections once the roots have grown so that each student or group of students doesn’t need a whole clove. They do however, need at least 3 roots each. Place each section of the clove back on a cocktail stick for ease of handling and a safer protocol.
You should use a garlic clove that has been growing for approximately 3 days with roots about 1cm long. The growth rate of roots is variable so it is worth setting up cloves over a series of a few days so that you have some roots of the right length available.
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