Patterns of resistance gene evolution in roses as models for heterozygous outcrossing woody perennials
Final Report Abstract
Black spot is the most severe disease on garden and field grown roses. The disease is caused by the hemibiotrophic fungus Diplocarpon rosae. Several isolates of Diplocarpon rosae are reported so far. In roses, the most extensively characterized gene that confers resistance to multiple isolates of Diplocarpon rosae is the Rdr1 gene. Using a map-based cloning approach the position of the Rdr1 gene was delimited to four overlapping BAC clones constructed from the resistant R. multiflora genotype. Complete sequencing and assembly of the four overlapping BAC clones carrying the Rdr1 gene resulted in a continuous sequence of 265477 bp. Annotation of the sequence revealed 47 predicted protein coding genes. In an attempt to understand the pattern of Rdr1 gene evolution, homologous regions from a susceptible rose cultivar were analysed. Hence, another four overlapping BAC clones constructed from the homologous region of the susceptible R. rugosa genotype were completely sequenced. A sequence length of 346835 bp and a similar number of protein coding genes were predicted for the homologous region. Nine and eleven TIR-NBS-LRR (TNL) type disease resistance genes were among those genes located on the R multiflora and R. rugosa BAC clones, respectively, considered potential candidate gene for Rdr1. Evolutionary analysis of these 20 Rdr1 candidate genes indicated the clusters evolved before the two species separated. In a comparison of these gehes to hundreds of Rdr1 homologues from other Rosaceae shows that the genes from Rosa cluster much closer than genes from other genera. This indicates that duplication of some of the Rdr1 genes occurred after genus Rosa diverged from other genera of the Rosaceae. Two of the Rdr1 candidate genes (muRdr1B and muRdr1G) are highly similar to each other with a nucleotide diversity p value of 0.0061 and without any indications of intragenic reshuffling, indicating recent duplication of at least these genes. In the course of sequence analysis an SSR marker (Rd1LRR) was developed from the coding region of the last exon of the Rdr1 candidate genes. SSR-marker results show a large variation in gene copy number among genotypes from the same species as well as between species. Gene duplication and deletion in the Rdr1 gene family is therefore a continuing process in Rosa regardless ofthe long-living perennial nature of roses. Our analyses of the ratio of synonymous versus non-synonymous substitutions revealed ratios of Ka/Ks <1, along the entire protein coding regions ofthe Rdr1 candidate genes. This indicates that positive selection for diversification is less frequent in perennial rose species as compared to other plant hosts which have been studied so far. Evolutionary mechanisms including illegitimate recombination, gene conversion, unequal crossing-over, indels, point mutation and transposable elements were identified, not principally different from that of short-living annual crop plants.
Publications
- (2008): Sequence-based comparison of the genetic region spanning the Rdr1 gene in two different rose species. XX International Congress of Genetics, July 12-17, 2008, Berlin
Terefe, D., Biber, A., Kaufman, H. and Debener, T.
- (2009): Sequence analyses of BAC contigs spanning the resistance gene Rdr1 against black spot (Diplocarpon rosae) in roses. Plant and Animal Genome XVII Conference, January 10-14, 2009, San Diego, .CA
Terefe, D. and Debener, T.
- (2010): An SSR from the leucine-rich repeat region of ttie rose Rdr1 gene family is a useful resistance gene analogue marker for roses and other Rosaceae. Plant Breeding
Terefe, D. and Debener, T.
(See online at https://doi.org/10.1111/j.1439-0523.2010.01780.x) - (2010): Comparative genomic analysis of sequences around the Rdr1 locus in resistant and susceptible rose genotypes. Acta Horticulturae
Terefe, D., Biber, A., Yasmin, A., Kaufmann, H., Linde M. and Debener, T.
- (2010): Dissecting rose NBS-LRR encoding genes by means of an exon based SSR marker. 10. GPZ Haupttagung, März 15-17, 2010, Freising-Weihenstephan
Terefe, D. and Debener, T.
- (2010): Identification and Characterization of the Rdr1 Resistance Gene from Roses. PhD Thesis, Leibniz University of Hannover
Yasmin, A.
- (2010): Molecular markers from a BAC contig spanning the Rdr1 locus: a tool for marker-assisted selection in roses. Theor Appl Genet. 120:. 765-773
Biber, A., Kaufmann, H., Linde, M., Terefe, D., Spiller, M. and Debener, T.
