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The soluble proteins in sieve tube exudate from Brassica napus plants were systematically analyzed by 1‐DE and high‐resolution 2‐DE, partial amino acid sequence determination by MS/MS, followed by database searches. 140 proteins could be identified by their high similarity to database sequences (135 from 2‐DE, 5 additional from 1‐DE). Most analyzed spots led to successful protein identifications, demonstrating that Brassica napus, a close relative of Arabidopsis thaliana, is a highly suitable model plant for phloem research. None of the identified proteins was formerly known to be present in Brassica napus phloem, but several proteins have been described in phloem sap of other species. The data, which is discussed with respect to possible physiological importance of the proteins in the phloem, further confirms and substantially extends earlier findings and uncovers the presence of new protein functions in the vascular system. For example, we found several formerly unknown phloem proteins that are potentially involved in signal generation and transport, e.g., proteins mediating calcium and G‐protein signaling, a set of RNA‐binding proteins, and FLOWERING LOCUS T (FT) and its twin sister that might be key components for the regulation of flowering time.