New partial sequences of phosphoenolpyruvate carboxylase as molecular phylogenetic markers.

To better understand the evolution of the enzyme phosphoenolpyruvate carboxylase (PEPC) and to test its versatility as a molecular character in phylogenetic and taxonomic studies, we have characterized and compared 70 new partial PEPC nucleotide and amino acid sequences (about 1100 bp of the 3' side of the gene) from 50 plant species (24 species of Bryophyta, 1 of Pteridophyta, and 25 of Spermatophyta). Together with previously published data, the new set of sequences allowed us to construct the up to now most complete phylogenetic tree of PEPC, where the PEPC sequences cluster according to both the taxonomic positions of the donor plants and the assumed specific function of the PEPC isoforms. Altogether, the study further strengthens the view that PEPC sequences can provide interesting information for the reconstruction of phylogenetic relations between organisms and metabolic pathways. To avoid confusion in future discussion, we propose a new nomenclature for the denotation of PEPC isoforms.

Molecular phylogeny of the genus Kalanchoe (Crassulaceae) inferred from nucleotide sequences of the ITS-1 and ITS-2 regions.

The study presents an analysis of genotypic diversity in the genus Kalanchoe (Crassulaceae) on the level of Internal Transcribed Spacer (ITS) sequences and the attempt to correlate this diversity with previous findings on ecophysiological behavior, habitat preference, infrageneric taxonomic position of the species and DNA polymorphism derived from RAPD-PCR data. The Kalanchoe species are mainly abundant in Madagascar and eastern continental Africa and perform in situ diverse modes of crassulacean acid metabolism (CAM), an ecophysiologically relevant adaptation of photosynthesis. Total DNA was extracted from 68 Kalanchoe species and varieties. The ITS-1 and ITS-2 regions of the nuclear RNA genes were amplified by polymerase chain reaction, cloned and sequenced. The alignments of the sequences were evaluated by distance (neighbor joining) and character state (maximum parsimony) methods. The main topologies of the obtained ITS phylogenetic trees were quite similar irrespective of the mode of evaluation and show: (A) within the Crassulaceae the genus Kalanchoe forms a monophyletic clade; and (B) within the genus the species form three main clusters which coincide well with the previously reported three infrageneric sections of the species distinguishable by classical taxonomic criteria, the mode of in situ CAM performance, and DNA fragment pattern obtained by RAPD-PCR analyses. Moreover, the ITS phylogenetic trees show that all African Kalanchoe species form a distinct group within the most derived of the three main clusters. This is consistent with the view that the center of phylogenetic radiation of the genus is located in Madagascar from where the species have spread into the continental Africa.