DNA systematic studies in the genus Bromus L. (Poaceae)

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Virginia Tech

Chloroplast and ribosomal DNA restriction endonuclease analyses were used to assess phylogenetic relationships between different subgenera of Bromus, a genus of over 100 diploid and polyploid species. Variation in chloroplast DNA (cpDNA) fragment pattern was examined, initially, in 15 species of subgenera Festucaria and Ceratochloa. Subsequently, cpDNA restriction site variation was examined in 38 species, using 10 enzymes and filter hybridization experiments. Variation in the ribosomal DNA (rDNA) repeat units was examined in 56 species, using four restriction endonucleases. Generally, higher polyploid species of Bromus showed very little or no interspecific variation in chloroplast DNA sequences. For example, nine species of Ceratochloa were identical in cpDNA structure. In contrast, diploid species showed various degrees of nucleotide sequence divergence. Cladistic analysis of cpDNA restriction site variation indicated two major lines of evolution within Bromus. One clade includes species of Festucaria, Neobromus and Ceratochloa and the other comprises species of Stenobromus and Bromus. cpDNA trees indicate greater genetic relationships among the subgenera of Bromus when compared to results from morphology and cytogenetics. The restriction site data suggests that Festucaria, Ceratochloa and Neobromus are closely related, while Stenobromus and Bromus are genetically isolated from the other subgenera and appear to be evolutionarily advanced. However, the cpDNA results cannot differentiate species of subgenera Stenobromus and Bromus since species of both subgenera were interwoven in the same clade. The cpDNA results suggest that additional characters are needed to provide further information on the taxonomy and evolution of Bromus. Restriction analysis of rDNA produced a wide variety of patterns. Digestion with BamHI produced extensive length heterogeneity within and among species. EcoRI digests were not useful for phylogenetic analysis since the enzyme generally cleaves only once per repeat unit. EcoRI and Kpni endonuclease fragment patterns were used to identify the number of repeat unit length variants per species. The BstEIIand Kpnl patterns were useful in determining relationships at the subgenus level. The unique 2.1 kb BstEII fragment of the coding region in subgenera Festucaria, Ceratochloa, Neobromus and Stenobromus suggests close genetic relationship among these subgenera. A similar fragment of 1.1 kb was present in subgenus Bromus suggesting genetic isolation from the other subgenera of Bromus.