Cytochemical investigations on multiple bud formation in tissue cultures of Pinus coulteri

Abstract

Cytochemical events associated with formation of multiple shoots on cultured embryonic explants of P. coulteri were studied. The 1st step in the developmental pattern leading to multiple adventitious bud formation was the establishment of organogenetic domains defined by increased RNA, DNA and basic nuclear proteins. The succeeeding step was the accumulation of reserves such as lipids, proteins and especially starch. This was followed by utilization of reserves characterized by high lipase and amylase activity in the domains. However, some of the subjacent callus cells also exhibited high lipase and amylase activity, indicating the mobilization of reserves for organogenesis. As meristemoids formed, there was an increase in enzyme activities concerned with energy production and utilization. Differentiating shoot primordia and developing needles showed high activities of acid phosphatase, glucose-6-phosphatase and succinate dehydrogenase. Intense localization of reducing substances (chiefly ascorbic acid) and increased ATPase activity suggested that there was a considerable utilization of energy associated with elevated metabolism in the meristemoids and developing buds. Peroxidase was strongly localized in regions of growth and differentiation, especially developing vascular strands. These cytochemical events apparently reflect distinctive changes in cellular metabolism associated with the initiation and development of multiple shoots on the cultured explants of P. coulteri.