Fine mapping of the SYM2 locus of pea linkage group 1

The symbiotic interaction between <em>Rhizobium</em> bacteria and leguminous plants results in the formation of root nodules which are specific, nitrogen-fixing organs that supply the plant with ammonia required for its growth. The formation of a nitrogen-fixing root nodule involves a complex series of steps requiring the expression of genes in both the rhizobial symbiont and the host plant. The necessary genes of rhizobia for nodulation, the <em>nod</em> genes, and for nitrogen fixation, the fix and <em>nif</em> genes, are well studied, nearly all of them having been cloned and characterized. In contrast the symbiosis genes of the legumes are not well understood. A number of plant genes which are specifically expressed in nodules or display enhanced expression in nodules, the so-called nodulin genes, have been identified in pea, soybean, alfalfa and other legumes by mRNA analysis and cDNA cloning. The time of expression of several nodulins has been analyzed in a number of cases and the nodulin genes which are expressed shortly after infection and in the first steps of nodule formation are referred to as early nodulins (ENODs).In addition, a large series of naturally occurring and induced plant mutants, the <em>sym</em> mutants, which have an altered symbiosis, have been described but the <em>sym</em> genes have not been characterized and the functions of the proteins encoded by these genes are not known. Since it seems a reasonable assumption that there will be a limited number of genes involved in nodule formation and metabolism it might well be that some of the <em>sym</em> mutants represent defects in nodulin-coding or controlling sequences. The study presented in this thesis is centred on one of the <em>sym</em> genes, <em>sym2,</em> as we suspected that the <em>sym2</em> gene has a role in the first interaction between the <em>Rhizobium</em> bacteria and the legume host plant. A typical characteristic of the <em>Rhizobium</em> legume symbiosis is the host specific nature. Most <em>Rhizobium</em> species can nodulate only plants of a specific plant genus. In the recently past years it has been demonstrated that in the <em>Rhizobium</em> bacteria the <em>nod</em> genes are responsible for the strong specificity in the bacteria-plant interaction. The <em>nod</em> genes are involved in the synthesis of specific lipo-oligosaccharides called Nod factors, that can induce the early responses in host plants leading to root nodule formation. Nod factors with a very specific structure are required to induce these responses and they are active at very low concentrations. Therefore it is probable that, as a first step in inducing the reactions leading to nodule formation, the Nod factors are recognized by a special receptor in the host plant. There are now several reasons to presume that the plant <em>sym2</em> gene might encode the receptor for the specific recognition of Nod factors. The aim of the study presented in this thesis is a further characterization of the interaction between <em>sym2</em> containing pea lines and different strains of <em>Rhizobium leguminosarium</em> bv. <em>viciae</em> in an attempt to find further support for this hypothesis. At the same time, we have started the detailed genetic mapping of <em>sym2</em> on the pea genome, in preparation for eventually cloning and molecular characterization of <em>sym2</em> .In chapter 1 a general introduction summarizes the history of the discovery of the <em>sym2</em> gene and presents the arguments for the hypothesis that <em>sym2</em> might code for a receptor of rhizobial Nod factors. In chapter 2 a further genetic characterization of <em>sym2</em> is given. Chapter 3 describes how a novel early nodulin gene of <em>PsENOD7</em> was characterized and mapped near the <em>sym2</em> locus. In chapter 4 a detailed map of the <em>sym2</em> locus linkage group I is presented including some new molecular markers tightly linked to the <em>sym2</em> locus. In chapter 5 it is demonstrated how the Rhizobium, <em>nodO</em> gene determines whether <em>sym2</em> is a dominant or a recessive allele. The thesis ends with some concluding remarks on the nature of <em>sym2</em> and the impact of the research described in the thesis for the cloning of <em>sym2</em> .