Sex Plant Reprod (1995) 8:254-255 9 Springer-Verlag 1995 Jo Oldknow 9 Tanya M. Franklin 9 Sharon Allard Laurian S. Robert 9 Martin Trick DNA sequences of the two homoeologous SLR1 genes of Brassica napus cv Westar Received: 10 April 1995 / Revision accepted: 18 April 1995 Key words Brassica napus 9 SLR1 genes 9 Pollination 9 Stigma-specific The Brassica SLR1 gene is related to the self-incompati- bility (SI) locus glycoprotein (SLG) gene that determines the specificity of pollen-stigma recognition (Nasrallah and Nasrallah 1993), although the function of the SLR1 protein is unknown. SLR1 genes are expressed in both SI and self-compatible (SC) Brassica lines (Lalonde et al. 1989; Trick and Flavell 1989) and in the related SC crucifer Arabidopsis thaliana (Lalonde et al. 1989; Dwyer et al. 1992). We report here DNA sequences for homoeologous SLR1 genes in the amphidiploid SC spe- cies Brassica napus. A genomic library was constructed from Brassica napus cv Westar in the bacteriophage vector )~FixII (Stratagene) and screened with an SLR1 cDNA probe (Trick and Flavell 1989). Several positively hybridising phages were isolated and found to correspond to overlap- ping regions around two homoeologous genes we desig- nated SLRlws ~ and SLRlws 2. Genomic sequences were corroborated with sequences of clones isolated from a stigma cDNA library. The DNA sequences of SLRlws 1 and SLRlws 2 are shown in Fig. 1. They predict proteins of 446 and 444 residues respectively, including N-terminal signal pep- tide sequences of 27 and 28 amino acids. The sequences share four putative N-glycosylation sites but SLRlws 2 has an additional site. Both predicted proteins conserve the 12 cysteine residues found in the C-terminus of all characterised SLG and SLR1 sequences. The two DNA sequences are 5% diverged, with 75% of the nucleotide The DNA sequence data reported have been lodged in the Gen- bank, EMBL and DDBJ databases under the accession numbers Z21609 and Z26914 J. Oldknow - T. M. Franklin 9 M. Trick (~) Department of Brassica and Oilseeds Research, John Innes Centre, Norwich Research Park, Colney, Norwich NR4 7UH, UK; Fax +44 (0) 1603 259882, e-mail: trick@bbsrc.ac.uk S. Allard 9 L. S. Robert Plant Research Centre, Central Experimental Farm, Ottawa, Ontario, Canada K1A 0C6 substitutions appearing in the N-terminal half. SLRlws ~ is most similar to the B. campestris NS I and NS 3 cDNAs (Isogai et al. 1991; Yamakawa et al. 1993), whereas SLRlws 2 is most like the SLR163 allele cloned from B. olereacea (Trick 1990). We have mapped the SLRlws 1 and SLRlws z genes to homoeologous linkage groups with precisely these diploid origins in the B. napus amphidiploid. RNA blot analysis indicated that SLR1 transcripts ap- peared only in maturing stigmatic tissue, peaking at 1 day pre-anthesis but still detectable 4 days after. This pattern of transcript abundance and developmental regu- lation is as previously demonstrated for both SLR1 and SLG genes in diploid Brassica oleracea. The extensive similarity between the homoeologous sequences preclud- ed a gene-specific transcript analysis. However, western blots of IEF gels of stigmatic proteins using a generic anti-SLR1 antibody showed the expression of two cross- reacting proteins migrating with the pIs predicted from the sequence data. The expression of active SLRI genes in both SC and SI plants suggests that the SLR1 protein may have some general role in pollen reception. We are attempting to define this function through the use of a gene ablation strategy. Aeknowleflgements We thank Dr Thierry Gaude for the SLR1 antibody. This work was supported by the AFRC PMB1 and EC BRIDGE programmes. References Dwyer KG, Lalonde BA, Nasrallah JB, Nasrallah ME (1992) Structure and expression of AtS1, an Arabidopsis thaliana gene homologous to the S-locus related genes of Brassica. Mol Gen Genet 231:442-448 Isogai A, Yamakawa S, Shiozawa H, Takayama S, Tanaka H, Kono T, Watanabe M, Hinata K, Suzuki A (1991) The cDNA sequence of NS 1 glycoprotein of Brassica campestris and its homology to S-locus-related glycoproteins of B. oleracea. Plant Mol Biol 17:269-271 Lalonde BA, Nasrallah ME, Dwyer KG, Chen C-H, Barlow B, Nasrallah JB (1989) A highly conserved Brassica gene with homology to the S-locus-specific glycoprotein structural gene. Plant Cell 1:249-258 Nasrallah JB, Nasrallah ME (1993) Pollen-stigma signalling in the sporophytic self-incompatibility response. Plant Cell 5:1325-1335