Abstract
We have used laser-capture microdissection to isolate RNA from discrete tissues of globular, heart, and torpedo stage embryos of Arabidopsis (Arabidopsis thaliana). This was amplified and analyzed by DNA microarray using the Affymetrix ATH1 GeneChip, representing approximately 22,800 Arabidopsis genes. Cluster analysis showed that spatial differences in gene expression were less significant than temporal differences. Time course analysis reveals the dynamics and complexity of gene expression in both apical and basal domains of the developing embryo, with several classes of synexpressed genes identifiable. The transition from globular to heart stage is associated in particular with an up-regulation of genes involved in cell cycle control, transcriptional regulation, and energetics and metabolism. The transition from heart to torpedo stage is associated with a repression of cell cycle genes and an up-regulation of genes encoding storage proteins, and pathways of cell growth, energy, and metabolism. The torpedo stage embryo shows strong functional differentiation in the root and cotyledon, as inferred from the classes of genes expressed in these tissues. The time course of expression of the essential EMBRYO-DEFECTIVE genes shows that most are expressed at unchanging levels across all stages of embryogenesis. We show how identified genes can be used to generate cell type-specific markers and promoter activities for future application in cell biology.
Highlights
We have used laser-capture microdissection to isolate RNA from discrete tissues of globular, heart, and torpedo stage embryos of Arabidopsis (Arabidopsis thaliana)
For an additional comparative time point, GeneChip data were utilized from the nonembryonic cotyledon and root tissue of a seedling, 7 d post germination. aRNA samples were shortened and showed a 3# bias (Supplemental Fig. S1)
These data are in a comparable range to these previous studies and partially bridge the temporal gap between them, demonstrating that at this arbitrary cutoff value a similar number of expressed genes are estimated throughout embryogenesis and in the mature Arabidopsis root
Summary
We have used laser-capture microdissection to isolate RNA from discrete tissues of globular, heart, and torpedo stage embryos of Arabidopsis (Arabidopsis thaliana). This was amplified and analyzed by DNA microarray using the Affymetrix ATH1 GeneChip, representing approximately 22,800 Arabidopsis genes. Time course analysis reveals the dynamics and complexity of gene expression in both apical and basal domains of the developing embryo, with several classes of synexpressed genes identifiable. The transition from globular to heart stage is associated in particular with an up-regulation of genes involved in cell cycle control, transcriptional regulation, and energetics and metabolism. The torpedo stage embryo shows strong functional differentiation in the root and cotyledon, as inferred from the classes of genes expressed in these tissues. Transcriptional Profiling of the Arabidopsis Embryo have since demonstrated the requirement for a larger number of essential genes based on the resulting frequency of embryonic lethality (Franzmann et al, 1995; McElver et al, 2001; Tzafrir et al, 2004)
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