Linking nucleotide polymorphisms in candidate genes and quantitative resistance to Melampsora larici-populina Kleb. in Populus nigra L

Association studies aim at linking phenotypic variation with allelic variation in candidate genes and benefit from several generations of recombination in natural populations to identify causative polymorphisms. Such association studies are under development on Populus nigra L., with emphasis on nucleotide polymorphism associated to quantitative resistance to Melampsora larici-populina Kleb (Mlp), the causal agent of leaf rust. Populus nigra, the natural host of Mlp, is widely represented in European riparian ecosystems, where the occurrence and impact of the fungus are highly variable. Geographical patterns of nucleotide diversity in genes involved in the host-pathogen interaction is expected from local co-evolution. As a first step, Single Nucleotide Polymorphisms (SNPs) have been identified in a discovery panel of twenty-one P. nigra genotypes covering French natural range. Sequencing was done on both functional and positional candidate resistance genes (closeness to mapped QTLs and resistance genes). An automated procedure for primer design allowed us to obtain successful amplification in P. nigra for 90% of gene fragments. Average nucleotide variation in P. nigra is high (1 SNP/97 pb). A collection of more than 1100 P. nigra genotypes collected in different natural populations mainly in France but also in Germany, Netherlands, Italy and Spain was evaluated for rust resistance under natural inoculation in a clonal test in France and under artificial inoculations with different strains in leaf-disk bioassays. Even if most of the genetic variation for rust resistant could be observed at the individual (genotype) level, significant differentiation between natural populations was detected. The high level of genetic diversity observed at both gene and phenotypic levels, the rapid linkage disequilibrium decay between SNP is offering a favourable situation to identify interesting resistance alleles and to avoid false positive associations.