A Modified Fuelgen Technique for
Staining the Chromosomes of Trilliums
L. Michael Hill
1. Obtain Fresh Root Tips.
2. Place the root tips into .2% colchicine for one hour at room temperature. Colchicine is a mitotic inhibitor that destroys the spindle fibers. This causes many metaphase plates to be seen because anaphase, which is the next stage of cell division, cannot happen because the fibers were destroyed. At metaphase, the chromosomes are at their most coiled state, easily seen, and easily sketched using the drawing tube of my microscope.
3. Fix the root tip in Carnoy's Fixative overnight or at 60 degrees Centigrade for 15 minutes.
4. Rinse in distilled water.
5. Hydrolyze in 1N Hydrochloric acid at 60 degrees centigrade for 12 minutes, no more no less. This is a critical step. The hyrdocholoric acid is reacting with the DNA and rendering it into a chemical state that will bind to the chemistry of the Fuelgen Stain. The temperature is critical, as is the timing.
6. Rinse in Distilled water.
7. Stain in Fuelgen Stain for 1-2 hours in the refrigerator.
8. Rinse in Distilled Water.
9. Separate slide from cover slip as per my instructions elsewhere in 45% acetic acid.
10. Recombine slide and cover slip in Euparal.
Sample results show considerable detail that can be measured, and one can also determine, with some accuracy, the morphology of the chromosome.
Source: Sharma, A.K., and A.S. Sharma. 1965. Chromosome Techniques, Theory and Practice. Butterworths, London.