This proposal requests funds for the development of novel flow cytometric instrumentation and operating procedures for use in the cytoenzymological analysis of large eukaryotic cells, those cells having diameters in the range of 20-l20 um. The research focuses on the development of novel substrates for use in flow cytoenzymology and on the development of techniques to permit the real-time analysis and sorting of cells based on surface area, volume, and concentration. The techniques will be employed for the analysis of those factors involved in the expression in plants of suitably engineered chimaeric genes, specifically beta- glucuronidase, following transfer from bacteria, and for the selection through flow sorting of living plant protoplasts according to the level of expression of the introduced genes.
