Viewing Mitosis in Onion Root Tips Lab 2 PDF

Lec.2 genetic Dr.Estabraq Almuhanna , Dr.Zainab AlKhalidi Viewing mitosis in onion root tips Requirements - Onion with roots - Plastic micro-tube (Eppendorf tubes) - 1N HCl (1 normal hydrochloric acid) - Water Bath - Distilled Water - Plastic pipette - Giemsa...

Lec.2 genetic Dr.Estabraq Almuhanna , Dr.Zainab AlKhalidi Viewing mitosis in onion root tips Requirements - Onion with roots - Plastic micro-tube (Eppendorf tubes) - 1N HCl (1 normal hydrochloric acid) - Water Bath - Distilled Water - Plastic pipette - Giemsa stain (DNA staining) - Microscope slide - lancet - Cover slip Procedure: First Step: The first step wigll be to ‘soften’ the roots, so they, later, can be spread on a microscope slide. 1. Using lancet, cut 2 roots tips about 1 cm long, and transfer them into a plastic micro-tube. 2. Fill the tube about 2 drop with 1N HCl by using pipette. *** Caution: Work with the HCl carefully, it is a strong acid. *** 3. Place the tube in a water bath, and allow the roots to incubate for 12 minutes. 4. After the 12 minute incubation period, remove the tube from the water bath. Reason: HCl and heat will soften the root tips. Lec.2 genetic Dr.Estabraq Almuhanna , Dr.Zainab AlKhalidi Second Step: Rinse the roots in H2O. 1. Using a plastic pipette, carefully remove the HCl from the micro-tube and transfer it to the “discard flask”. 2. Rinse the root tips 3 times with distilled water, and dispose disposing of the rinses in the discard flask. Reason: HCl must be removed, so it will not interfere with staining Third Step: Staining the chromosomes 1. After removing the water from the third rinse, cover the root with the Giemsa stain. 2. Incubate the roots in the stain for 12 minutes. as the DNA stains the numerous small actively dividing cells at the tip. Fourth Step: Remove the stain and again rinse the roots again. 1. Using a plastic pasture pipette, carefully remove stain) 2. Again, rinse the root tips 3 times with water. Fifth Step: Prepare the root tip. 1. Transfer a root to the center of a clean microscope slide and add a drop of water. 2. Using a lancet, cut off most of the unstained them. 3. Cover the root tip with a cover slip Lec.2 genetic Dr.Estabraq Almuhanna , Dr.Zainab AlKhalidi Observations of onion root tip. Scan the slide under the 10x objective lens. Look for the region that has large nuclei relative to the size of the cell and among these cells, cells displaying stages of mitosis will be found. Switch to the 40X objective to make closer observations.