Long range PCR to study the ribosomal proteins genetic region as a phylogenetic marker for Frankia strains.. Frankia phylogeny

Taxonomic assignment of uncharacterized isolates to known species is a challenge that is generally met by sequencing the 16S rRNA genes or other conserved markers known to cause distortion due in part to their short length. We targeted the 30kb ribosomal proteins region, amplified it with specific primers and a long range PCR approach and obtained a phylogenetic reconstruction. All known clusters were recovered with this approach with a very high confidence and the topology was found to be highly dependent on the outgroup used, due to a long-branch attraction effect. The robust topology recovered was found to yield the asymbiotic cluster4 as the ancestor of all Frankia lineages, from which then emerged the generally obligate cluster2, then the other clusters (1 and 3). The strongly symbiotic cluster2 was also found to have the fastest and the asymbiotic cluster4 had the slowest mutation rate.