Polymerase chain reaction – restriction fragment length polymorphisms for assessing and increasing biodiversity of Frankia culture collections

Canadian Journal of Botany (Impact Factor: 1.4). 12/1999; 77:1261-1269. DOI: 10.1139/b99-083

ABSTRACT During the last few years, some Frankia culture collections that maintained a large number of unidentified and uncharacterized Frankia strains were closed because of funding shortages. To reduce the costs of maintenance, we evaluated the biodiversity of half of the Frankia strains from our collection, by polymerase chain reaction – restriction fragment length polymorphisms (PCR-RFLPs) of nifD–nifK intergenic spacer and 16S–23S rDNA intergenic spacer regions. In this way we were able to reduce the number of strains without reducing the biodiversity of the whole collection. In general the nifD–nifK target proved to be more polymorphic than the rrn target. From 51 isolates of Elaeagnus frankiae, PCR-RFLP results allowed us to detect 13 identical strains, and to predict that the genomic species P8 of Akimov and Dobritsa (1992) very likely agrees with genomic species 5 of Fernandez et al. (1989). Moreover, we revealed genomic groups not yet described, as well as intraspecific variability. For Alnus frankiae, the polymorphisms shown by both the nif and the rrn PCR-RFLPs revealed three host plant species-specific subgroups inside Frankia alni. An expandable data base was created to serve as reference for future biodiversity evaluations on both culture collections and unisolated Frankia populations. It will be accessible by Internet at the International Frankia Website ( Résumé : Ces dernières années, plusieurs collection de cultures de Frankia contenant un grand nombre de souches non identifiées et non caracterisées ont été abandonnées par manque de fonds. Dans le but d'obtenir une réduction des coûts de maintien de notre collection, nous avons évalué la biodiversité existante dans la moitié des nos souches de Frankia. En analysant par la méthode réaction en chaîne de la polymérase – polymorphismes de la longeur des fragments de restriction (PCR-RFPLs) les régions intergéniques nifD–nifK et rrn 16S–23S, nous avons pu reduire le nombre de souches en culture, sans réduire la biodiversité de la collection. En général, l'étude de la région nifD–nifK a donné plus de polymorphisme que celle de la région ribosomale. Entre les 51 Frankia isolés d'Elaeagnus, on a pu détecter 13 souches identiques et montrer que les espèces génomiques P8 de Akimov et Dobritsa (1992) et 5 de Fernandez et al. (1989) pourraient être une même espèce. En plus, on a détecté des groupes génomiques pas encore décrits et une certaine diversité intra-spécifique. Pour les souches isolées d'Alnus, les polymorphismes montrés par les deux regions génomiques ont permis de mettre en evidence, chez espèce Frankia alni, trois sous-groupes spécifiques pour chaque espèce hôte. Pour faciliter les travaux futurs sur la biodiversité des Frankia cultivés ou des populations non isolées, on a construit une banque de profils PCR-RFLP qui sera accessible via l'Internet dans le site Web International sur Frankia (http:

  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The capacity of some Tunisian soils to induce nodulation on Casuarina spp. and Alnus glutinosa was investigated through survey at fields and by plant-trapping bioassay. Frankia nodules were detected only in the north of Tunisia in some experimental forest sta-tions for Casuarinaceae, and in natural endemic A. glutinosa stands. Frankia genetic diver-sity was assessed by PCR-RFLP of nifD-K region and, for Casuarinaceae, also of 16S-23S rDNA internal transcribed spacers, amplified from DNA directly extracted from root nod-ules. Restriction patterns showed that one and two haplotypes of Frankia colonise Casuar-inaceae and A. glutinosa, respectively. Frankia in nodules of Casuarinaceae were found to be closely related to the group 1 of Casuarinaceae nodulating strains previously identified in Australia, corroborating the hypothesis of a recent introduction of these strains into Tunisia, probably with their hosts.
    Annals of Microbiology 01/2002; 52:145-153. · 1.04 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Symbiotic association of the N 2 -fixing actinomycete Frankia with the roots of more than 200 tree species from 24 genera of 8 families of angiosperms has been studied since 1829. The first successful isolation of the microsymbiont and reinfection in the host plant was achieved in 1978. Marked advances in research and understanding of Frankia biology, its actinorhizal hosts and their interactions have been made since then, although the studies on Frankia have been hampered by difficulties of isolation and their slow growth rate in vitro. Exponential growth with high biomass yields within three to four days has been obtained for a number of strains isolated from Casuarina spp. Use of BAP medium, supplemented with avian phospholipid mixtures and certain fatty acids at controled O 2 access, optimizes growth. Monosporal cultures are scarce; recently a few became available for biochemical and genetic studies. Research using exponentially growing cultures has yielded information on a complex proteolytic system, including proteasomes, endo- and extracellular proteinases and aminopeptidases, and also on esterases, dehydrogenases, and extracellular DNAses. Molecular tools have revealed a marked genetic diversity of Frankia soil populations and have enabled the definition of four clades in the Frankia phylogenetic tree. Studies on Frankia -host plant interactions have detected molecular signal exchange preceeding the establishment of symbiosis. Similarly, there is progress in research on transgenic actinorhizal plants and on actinorhizal-specific genes and proteins (actinorhizins) involved in symbiotic interactions, infectivity, and host specificity. Actinorhizal plants are rapidly growing species, able to develop in N-poor soils, and for certain species, in harsh environmental stress conditions. They increase the fertility of agroforestry ecosystems, and have an economic potential for timber, fuelwood production, land reclamation, and amenity planting. The Casuarina spp. are of especial value in arid environments.
    Arid Land Research and Management 10/2001; 15(4):285-327. DOI:10.1080/153249801753127615 · 0.56 Impact Factor
  • Source
    African journal of microbiology research 02/2014; 8(20):2006-2010. DOI:10.5897/AJMR2014.6826 · 0.54 Impact Factor


Available from
Jun 3, 2014