Analysis of labeling of plant protoplast surface by fluorophore-conjugated lectins

Abstract

Figures 3 and 8 should be changed over. The lettering of the abscissa of Fig. 3 should be read "Drug concentration (gg/ml)" instead of "Drug concentration (ug/ml)".

Mosaicism in the organization of Con A binding sites on the membrane surface of female cells of Nicotiana tabacum

“…3, panel 3b). As previously reported, enzymatic treatment did not noticeably change the distribution of lectin binding sites (Burgess & Linstead, 1976;Walko et al ., 1987;Fang et al ., 2003). Furthermore, some egg cells and central cells were placed in 1% paraformaldehyde dissolved in 9% mannitol for fast surface fixation immediately after isolation.…”

“…Thus, binding was specifically identified in our competitive inhibition experiments. In another control experiment, the plasma membranes of somatic protoplasts exhibited a smooth, uniform fluorescent ring that was observable from any orientation, as previously described by Walko et al . (1987).…”

“…Tobacco plant ovules were dissected from ovaries and placed into an enzyme solution composed of 13% mannitol, 1% cellulase R-10 and 0.8% macerozyme R-10, pH 5.7, and incubated for 3 h with agitation. The enzymes have been tested and confirmed that they do not influence the distribution and binding ability of lectin binding sites (Burgess & Linstead, 1976;Walko et al ., 1987;Sun et al ., 2002a;Fang et al ., 2003). Three washes in 13% mannitol were then performed to remove any remaining enzymes.…”

“…Binding specificity test Binding specificity was tested using the procedures described above, except that the incubation medium contained competing monosaccharides, primarily α -methyl-D-mannoside. In this case, the materials were preincubated with 0.1 M α -methyl-D-mannoside for at least 30 min before the cells were labeled (Walko et al ., 1987).…”

A family of abundant plasma membrane-associated glycoproteins related to the arabinogalactan proteins is unique to flowering plants.

“…Agglutination and fluorescence labeling of plant protoplasts by lectins has permitted the identification of terminal glycan residues on the outer face of the plasma membrane (Walko et al, 1987), presumably components of glycoproteins or glycolipids. The presence among these of members of the class of glycoproteins termed arabinogalactan proteins (AGPs)' (Fincher et al, 1983) has been surmised from protoplast agglutination with the/3-glycosyl Yariv reagents (Larkin, 1977(Larkin, , 1978Samson et al, 1983), and from the coincident migration during isoelectric focussing of hydroxyproline-rich membrane components and components with affinity for ~-galactosyl Yariv reagent (Samson et al, 1983).…”