FLUORESCENT IMAGING OF GUS ACTIVITY AND RT-PCR ANALYSIS OF GENE-EXPRESSION IN THE SHOOT APICAL MERISTEM

The use of promoter-reporter gene constructs in transgenic plants is a powerful tool in the analysis of gene expression which can, however, be limited in the resolution of small structures, such as the apical m eristem. This paper reports on a fluorescent imaging technique for the analysis of GUS reporter gene expression to cellular resolution in th e apical meristem of tomato. Using this technique in combination with an RT-PCR analysis of RES gene-specific transcript levels, it is shown that: 5' upstream sequences of RBCS genes are sufficient to mimic the pattern of transcripts revealed by in situ hybridisation (no expressi on in the apical meristem, high expression in the initiated leaf primo rdial; the genes RBCS2, RBCS3A and RBCS3B are transcriptionally activa ted upon primordium initiation with transcripts for RBCS1 and RBCS3C a ccumulating later in leaf development; and that RBCS promoter activity cannot be induced in the apical meristem by light, an environmental s ignal which elevates RBCS transcript level in other aerial parts of th e plant. These data provide a detailed picture of the complex transcri ptional events occurring on leaf initiation and the establishment of t he photosynthetic machinery; they describe two complementary technique s which allow the analysis of such complex events at the tissue and ce llular level; and they characterize an in vivo assay system which can be used to analyse the factors involved in the initiation and maintena nce of gene expression patterns in the apical meristem.