The diversity, ecology and phylogeny of Frankia in actinorhizal plants

Date of Completion

January 1999


Biology, Ecology|Biology, Microbiology




Nitrogen-fixing symbiotic strains of the genus Frankia were characterized in field collected angiosperm (actinorhizal) root nodules. The nodules were dissected under a microscope and symbiont genomic DNA was purified using standard alkaline lysis ammonium acetate/ethanol precipitation. Either full or partial 16S rRNA as well as Glutamine Synthetase (glnA) genes were amplified using appropriate PCR primers. The amplicons were purified on DNA columns, quantified and sequenced. Approximately 250 genes were amplified and sequenced. The sequences were aligned on a VAX computer using the GCG package and ClustalW. The alignments were used to construct phylogenetic trees with various programs (PHYLIP, PUZZLE) and algorithms (Kitsch, Maximum likelihood, Neighbor-Joining, Parsimony, UPGMA). The 16S rRNA genes were also used as ecological markers to assess the diversity of Frankia symbionts their actinorhizal hosts. The richness of Frankia 16S rRNA genes in some plants was extrapolated using non-parameteric estimates (Abundance Coverage Estimator, Second Order Jackknife). The evolution of Frankia and actinorhizal plants was compared using Frankia 16S rRNA and glnA genes, and published actinorhizal chloroplast rbcL gene sequences. The notion that the actinorhizal symbiosis has multiple origins is supported by this study. The diversity of Frankia microsymbionts in actinorhizal hosts is variable, and examined in the context of an ecological mosaic. ^