A simple method for the identification and assay of extracellular plant beta-galactosidase.
A simple, rapid and reproducible procedure for the identification of extracellular Californian poppy (Eschscholzia californica Cham.) beta-galactosidase is described using callus cultures of seedlings from the tested plant, roots of 4-days-old seedlings of Californian poppy germinating on agar plates and cell suspension cultures cultivated from callus cultures. 6-Bromo-2-naphthyl-beta-D-galactopyranoside and p-nitrophenyl-beta-D-galactopyranoside were used as substrates for the determination of the intracellular and extracellular activities of beta-galactosidase. The extracellular beta-galactosidase activity was identified by evaluating the dye-zones in an agar medium. The enzyme from Californian poppy callus cultures or from seedling roots cultivated on agar plates supplemented with 6-bromo-2-naphthyl-galactopyranoside hydrolyzed this substrate releasing 6-bromo-2-naphthol. By simultaneous coupling with hexazonium p-rosaniline the corresponding (reddish-brown) azo-dye was formed. The agar plate method described permits rapid, simple and specific detection of plant producers of extracellular beta-galactosidase.
Stano
J
J
Faculty of Pharmacy, Department of Botany, Faculty of Sciences, Comenius University, Bratislava, Slovak Republic.
Mićieta
K
K
Neubert
K
K
Luckner
M
M
Adam
H
H
D002478:Cells, Cultured; D005110:Extracellular Space / Q000201:enzymology; D005697:Galactosides / Q000737:chemistry; D007202:Indicators and Reagents; D010207:Papaver / Q000737:chemistry; D010944:Plants / Q000201:enzymology*; D015203:Reproducibility of Results; D001616:beta-Galactosidase / Q000032:analysis*
D005697:Galactosides; D007202:Indicators and Reagents; C065297:6-bromo-2-naphthyl-beta-galactopyranoside; D001616:beta-Galactosidase