Histology of, and physical factors affecting, transient GUS expression in pearl millet (Pennisetum glaucum (L.) R. Br.) embryos following microprojectile bombardment.
Transient GUS (β-glucuronidase) expression was visualized in whole and sectioned embryos of Pennisetum glaucum (L.) R. Br. (pearl millet) after microprojectile bombardment with pMON 8678 DNA. Strongest GUS expression occurred in cells located in the center of GUS positive spots with decreasing intensity in surrounding cells. GUS positive cells could be seen up to 12 cell layers beneath the epidermis. Needle-like crystals of the GUS assay product were found throughout the cytoplasm of GUS positive cells. The number of GUS positive spots was correlated to the microprojectile spread pattern on the medium surface. Shorter bombardment distances (6.6 and 9.8 cm) and the standard accelerator speed gave the best results for transient expression but also caused maximum tissue damage. The speed and distance, however, had little influence on the ability of bombarded embryos to form compact callus. The developmental stage of the bombarded immature embryos was the determining factor in the formation of compact callus, from which plants were regenerated.
Laboratory of Plant Cell and Molecular Biology, Department of Vegetable Crops, University of Florida, 32611, Gainesville, FL, USA.
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Vasil