Multiple independent losses of the plastid rpoC1 intron in Medicago (Fabaceae) as inferred from phylogenetic analyses of nuclear ribosomal DNA internal transcribed spacer sequences

A previous polymerase chain reaction based survey for the occurrence of the intron in chloroplast gene rpoC1 revealed its absence in one of the eight species of Medicago (Fabaceae; Trifolieae) examined. We extend the survey of Medicago to include 65 of the 86 species, representing all 12 sections and seven of the eight subsections recognized in the most recent comprehensive treatment of the genus. Our results indicate that 17 species from five sections lack the intron and that three of these sections are heterogeneous with regard to intron content. DNA sequencing across the rpoC1 intron-exon boundary in three of these species reveals the precise excision of the intron from the gene. Phylogenies derived from nuclear ribosomal DNA internal transcribed spacer sequences, estimated using maximum parsimony and maximum likelihood methods, suggest that most of the previously recognized sections in Medicago are not monophyletic as currently circumscribed. Furthermore, these results suggest that the rpoC1 intron has been lost independently a minimum of three times during the evolution of the group. The occurrence of multiple independent intron losses severely reduces the utility of this character as a phylogenetic marker in Medicago.