Seasonal changes in fluorescence intensity of kiwifruit nuclei stained with propidium iodide and measured by flow cytometry

Abstract

Nuclei were extracted from kiwifruit (Actinidia deliciosa) axillary buds and shoot apices at intervals during two growing seasons. Nuclei from fruit tissues were also examined. Extracted nuclei were stained with propidium iodide, which intercalates into double stranded nucleic acids, and the intensity of the staining reaction (fluorescence channel number) was measured by flow cytometry.

Fluorescence intensity (mean channel number) of 2C nuclei varied during seasonal growth in both 1991 and 1992. In 1991, channel numbers of nuclei from shoot apices of pistillate and staminate vines were highest at the time of bud break and then declined. A similar change was observed in nuclei from pistillate shoots grown in a glasshouse. In 1992, however, nuclei extracted during bud break from axillary buds and shoot apices had low channel numbers. In this year, highest channel number values occurred during mid summer and the values for nuclei from fruit tissues were also high at this time. Lower channel number values were obtained in late summer for both axillary buds and fruit tissues.

When nuclei with low channel numbers were treated with 0.1N hydrochloric acid for 1.0–1.5 min, channel numbers were increased. In 1991, the acid treatment fully restored channel numbers to maximum values, but, in 1992, complete restoration only occurred for nuclei sampled during late winter and early spring.

Changes observed in channel number values of nuclei from fresh tissues most likely resulted from conformation changes in the nucleus which altered the intercalation of propidium iodide. However, altered propidium iodide intercalation did not fully explain lowered channel numbers during spring and summer in 1992. Changes found in the nuclei are discussed in relation to periods of cell differentiation in shoot and fruit tissues.

Keywords:

• kiwifruit
• nuclei
• propidium iodide
• flow cytometry
• fluorescence intensity
• seasonal changes