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The pea mutant line P55 is defective in root nodule formation, and this phenotype is controlled by a single recessive gene. Complementation analysis revealed that the mutation in P55 is allelic to sym19, which has previously been mapped to linkage group I. Detailed mapping revealed that the sym19 and ENOD40 loci are separated by 2.7 cM. We identified four recombination events, demonstrating that the nodulation defect caused by mutation of the sym19 locus cannot be due to mutation of ENOD40. RT-PCR experiments showed that P55 expresses ENOD12A, but there was little or no increase in the level of its transcript in response to Nod factor or infection with Rhizobium. To investigate this expression pattern further, transgenic peas carrying a pENOD12A-GUS reporter construct were made. One transgenic line was crossed with line P55, to generate F2 progeny homozygous for sym19 and carrying pENOD12A-GUS. In both WT and sym19 mutant lines, ENOD12A-GUS expression was induced at sites of lateral root emergence in uninoculated plants. In Nod+ plants pENOD12A-GUS was induced in response to Rhizobium leguminosarumn bv. viciae, but no such induction was seen in the Nod- (sym19) mutants.

Within a set of 13 induced symbiotic pea (Pisum sativum L.) mutants, two distinct mutated genes sym25 and sym26, selected for ineffective modulation have been counteracted in their effect by Rhizobium strains 254 and 94 respectively, thereby establishing an N2-fixing symbiosis. This suggests that mutagenesis can be a helpful tool in creating specificity systems such as those described in centres of origin and diversification of legume species. [ABSTRACT FROM AUTHOR]