A cytogenetic study of factors affecting sister chromatid differentiation in Vicia faba and Hordeum vulgare.

Abstract

A cytological study has been carried out with Vicia faba and Hordeum vulgare to examine a number of techniques for diferentially visualizing sister chromatid exchanges (SCE) using the Fluoresent-plus-Giemsa and the Feulgen procedures. Factors which affect seedling growth and BrdU uptake of growing seedlings are important for sister chromatid differentiation. A variety of steps carried out after the root tips are harvested, including fixation and staining have been shown to affect both the quality of staining and differentiation of SCE. Other important factors to be considered if this technique is to be used in mutagenicity testing are the concentration of both BrdU and FdU, and the degree of squashing of the cells to obtain good metaphase spreads for visualizing the chromosomes. Treatments such as the removal of plant shoots and cotyledons, and the application of BrdU and FdU led to reductions in mitotic indices. However, it is not recommended to eliminated FdU, since it has been shown to enhance the uptake of radioactively labelled BrdU. A concentration of 100μM BrdU was satisfactory when root tips of whole plants were treated. The variance of the mitotic index within treatments when seedlings were treated was consistently high, implying that a range of tolerance to a given treatment existed among individual plants of the same cultivar. This variance caused the results of statistical analyses to be inconclusive with respect to differences between treatments. Fixation had a significant influence on the quality of staining and a fixation time of 7 hours is suggested. Fixation should be carried out at 4°C and storage of stained root tips for up to one month in 70% ethanol does not seem to adversely affect staining. Good differential staining was obtained in Vicia and in barley by means of the Feulgen procedure, but not the FPG technique.