Electron microscopy and X-ray microanalysis as tools for fine localization of the β-glucuronidase activity in transgenic plants harbouring the GUS reporter gene

摘要: The GUS reporter gene encoding β-glucuronidase is very useful in various domains of plant genetic engineering. A method for ultrastructural detection its activity was developed using 35S-GUS transgenic tobacco root tips. Short glutaraldehyde prefixation at 4°C preserved up to 70% enzyme and followed by brief incubation X-Glu, strong postfixations, then quick dehydration low temperature before resin embedding. In these conditions, cells were well displayed electron dense indigo precipitates with a crystalline structure as shown diffraction. Due other structures the tissues, controls nature reaction product (diX-indigo) necessary. first control carried out means X-ray microanalysis order check presence bromine. Other controls, including incubated non-transformed non-incubated or boiled roots samples specific inhibitor, D-saccharic acid-1,4-lactone, also out. discussion points potential uses but limits method, non-specific localizations diX-indigo microcrystals being possible.