Structural, physiological and biochemical aspects of embryo development in vivo and in vitro

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Apical meristems are located at each growing point and are specialized for the production of new cells for the plant body.  The importance of meristems in plant growth and development can be summarized by a simple statement from Professor Ian Sussex's review "The meristems make the plant" (Cell 56: 225, 1989). In the past funding period, we demonstrated that shoot meristem initiation is sensitive to auxin perturbation.  Furthermore, during in vitro embryogenesis, although a shoot meristem is formed, it is not stably determined as its organization deteriorates upon prolong culture. The root meristem, on the other hand, requires auxin for development. These observations suggest that expression of the meristem genes and hence the cell fate could be altered by auxin levels and factors from the embryonic environment. We also established that the redox state of both the cell and its surrounding environment can modulate in vitro embryo development.  The redox environment can influence the quality of the embryos produced, especially the shoot apical meristem.There are still many important, unanswered questions regarding meristem initiation and its determination process.  At present, we have in our collection a number of Arabidopsis meristem-related enhancer-trap lines with GUS (beta-glucuronidase) as the marker gene.  In the next funding period, we will make use of these enhancer-trap lines to study meristem formation and the role of auxin and other medium additives during in vitro development.  The advantage of using the GUS marker gene is that once this gene is subsequently translated, we can stain for the enzyme activity which gives a blue color.  By studying changes in the color pattern, we can study the activities of various meristem genes during the course of meristem development.  We will further investigate the importance of the redox environment on embryo development.  The white spruce system will be used. The role of the redox compounds, i.e. reduced and oxidized glutathione on embryogenic tissue proliferation and programmed cell death will be studied using histological, histochemical, and biochemical methods.