Molecular and morphological phylogenetic analysis of Brachiaria and Urochloa (Poaceae).

The taxonomic relationships of Brachiaria and Urochloa have been questioned based on previous morphological studies. In this paper, we reconsider the phylogenetic relationships of these genera using 22 species of Brachiaria and Urochloa and six species of Paniceae as out-groups. The ITS1, 5.8S, and ITS2 region (internal transcribed spacer) of nuclear ribosomal DNA and eight morphological characters of the inflorescence were compiled into a data matrix. The cladistic analyses suggest that Urochloa-Brachiaria as a complex is paraphyletic with Eriochloa and Melinis. Species of all these genera share molecular synapomorphies and belong to the same monophyletic groups. The results confirm the continuous gradation between those genera previously found in several morphological studies. Therefore, the following eight new combinations are made: Urochloa bovonei (Chiov.) A.M. Torres & C.M. Morton, Urochloa dura (Stapf) A.M. Torres & C.M. Morton, Urochloa dura var. dura (Stapf) A.M. Torres & C.M. Morton, Urochloa dura var. pilosa (J.G. Anderson) A.M. Torres & C.M. Morton, Urochloa lachnantha (Hochst.) A.M. Torres & C.M. Morton, Urochloa leersioides (Hochst.) A.M. Torres & C.M. Morton, Urochloa nigropedata (Munro ex Ficalho & Hiern) A.M. Torres & C.M. Morton, and Urochloa subulifolia (Mez) A.M. Torres & C.M. Morton.

An intraspecific genetic map of velvetbean (Mucuna sp.) based on AFLP markers.

Velvetbean ( Mucuna sp., n=11), a self-pollinated species, is an important legume used in tropical agricultural systems in rotation with other crops for nematode management and/or soil improvement. A genetic map of velvetbean was constructed in order to identify potential molecular markers linked to important morphological and agronomic traits that would be particularly useful for developing and improving the species. Traits such as seed coat color, pod color, and pod pubescence were among the main parameters observed in a process of genetic diversity estimation. Two slightly divergent velvetbean accessions, PI364362 and 'Edgar Farm White', a land race from Alabama, were used to make an intraspecific F(1) hybrid. Amplified fragment length polymorphism analysis (AFLP) detected an average of six polymorphic fragments per primer pair between the two parents. As expected for dominant markers, the sum of all AFLP bands from both parents was generally observed to be present in the AFLP profiles of the F(1) progeny, indicating full penetrance and the dominant nature of AFLP markers. An F(2) population was generated by self-pollinating a single F(1) plant. Using 37 AFLP primer pairs, we detected 233 polymorphic markers of which 164 (70.4%) segregated in 3:1 Mendelian ratios, while the remaining 69 (29.6%) both segregated and were scorable. The genetic linkage map constructed from this population comprised 166 markers, including two morphological traits (pod color and pod pubescence). Twenty linkage groups were found with an average distance between markers of 34.4 cM, covering a total of 687.9 cM. The linkage groups contained from 2 to 12 loci each and the distance between two consecutive loci ranged from 0 to 21.8 cM. The newly designated morphological traits pod color ( pdc) and pod pubescence ( pdp) co-segregated with each other at a distance of 4.2 cM. Two DNA markers designated ACGCAG2 and ACTCTG1 were located in the same group as pdc and pdp. The AFLP linkage map provides opportunities for use in marker-assisted selection and in the detection of loci controlling morphologically important traits.

Transcriptional regulation of the mosquito vitellogenin gene via a blood meal-triggered cascade.

In anautogenous mosquitoes, a blood meal is required for activation of genes encoding yolk protein precursors (YPP). Vitellogenin (Vg), the major YPP gene, is transcribed at a very high level following blood meal activation. It is expressed exclusively in the female fat body, the tissue producing most of mosquito hemolymph and immune proteins. In this paper, we analyzed the upstream region of the Aedes aegypti Vg gene in order to identify regulatory elements responsible for its unique expression pattern. To achieve this goal, we analyzed the gene using transgenic Drosophila and Aedes as well as DNA-binding assays. These analyses revealed three regulatory regions in the 2.1 kb upstream portion of the Vg gene. The proximal region containing binding sites to EcR/USP, GATA, C/EBP and HNF3/fkh is required for the correct tissue- and stage-specific expression at a low level. The median region carrying sites for early ecdysone response factors E74 and E75 is responsible for hormonal enhancement of Vg expression. Finally, the distal GATA-rich region is necessary for extremely high expression levels characteristic of the Vg gene. The present work elucidates the molecular basis of blood meal-dependent expression of this mosquito gene, laying the foundation for mosquito-specific expression cassettes with predictable stage and tissue specificity.

Phylogenetics of flowering plants based on combined analysis of plastid atpB and rbcL gene sequences.

Following (1) the large-scale molecular phylogeny of seed plants based on plastid rbcL gene sequences (published in 1993 by Chase et al., Ann. Missouri Bot. Gard. 80:528-580) and (2) the 18S nuclear phylogeny of flowering plants (published in 1997 by Soltis et al., Ann. Missouri Bot. Gard. 84:1-49), we present a phylogenetic analysis of flowering plants based on a second plastid gene, atpB, analyzed separately and in combination with rbcL sequences for 357 taxa. Despite some discrepancies, the atpB-based phylogenetic trees were highly congruent with those derived from the analysis of rbcL and 18S rDNA, and the combination of atpB and rbcL DNA sequences (comprising approximately 3000 base pairs) produced increased bootstrap support for many major sets of taxa. The angiosperms are divided into two major groups: noneudicots with inaperturate or uniaperturate pollen (monocots plus Laurales, Magnoliales, Piperales, Ceratophyllales, and Amborellaceae-Nymphaeaceae-Illiciaceae) and the eudicots with triaperturate pollen (particularly asterids and rosids). Based on rbcL alone and atpB/rbcL combined, the noneudicots (excluding Ceratophyllum) are monophyletic, whereas in the atpB trees they form a grade. Ceratophyllum is sister to the rest of angiosperms with rbcL alone and in the combined atpB/rbcL analysis, whereas with atpB alone, Amborellaceae, Nymphaeaceae, and Illiciaceae/Schisandraceae form a grade at the base of the angiosperms. The phylogenetic information at each codon position and the different types of substitutions (observed transitions and transversions in the trees vs. pairwise comparisons) were examined; taking into account their respective consistency and retention indices, we demonstrate that third-codon positions and transitions are the most useful characters in these phylogenetic reconstructions. This study further demonstrates that phylogenetic analysis of large matrices is feasible.

Phylogenetic relationships of Rutaceae: a cladistic analysis of the subfamilies using evidence from RBC and ATP sequence variation.

Sequence data for plastid rbcL and atpB from members of Anacardiaceae, Burseraceae, Cneoraceae, Meliaceae, Ptaeroxylaceae, Rutaceae, and Simaroubaceae were analyzed cladistically to evaluate the familial and subfamilial circumscriptions of Rutaceae. Taxa representing all subfamilies and tribes were sampled. The analysis shows that Rutaceae are paraphyletic, with Spathelia and Dictyoloma (Rutaceae), Harrisonia (Simaroubaceae), Cneorum (Cneoraceae), and Ptaeroxylon (Ptaeroxylaceae) forming a clade sister to all other Rutaceae. Circumscription of Rutaceae to include all of these taxa is recommended. This analysis indicates that Simaroubaceae and Meliaceae are the outgroups closest to Rutaceae. Correlation of the molecular phylogenies with biochemical data indicates that chemotaxonomic information is more reliable than fruit type as an indicator of familial and subfamilial circumscriptions. The subfamilial classification needs revision; none of the subfamilies of more than one genus is monophyletic.

Biogeographic patterns in Mediterranean and Macaronesian species of Saxifraga (Saxifragaceae) inferred from phylogenetic analyses of ITS sequences.

A biogeographic study of Saxifraga section Saxifraga was performed based on phylogenetic analyses of ITS (internal transcribed spacer) sequences of nuclear ribosomal DNA. ITS sequences from 21 species and 31 populations were examined to identify colonization patterns for the two species of Saxifraga occurring in Macaronesia and for S. globulifera in the west Mediterranean basin. Phylogenetic analysis of the sequence data yield a single most parsimonious tree with many of the major clades well supported by bootstrap and decay values. The ITS tree provided resolution at specific and populational levels that points to two biogeographic patterns within the genus. In contrast to the molecular evidence provided by other authors for a Mediterranean origin of several Macaronesian genera of angiosperms, our results indicate that the Madeiran archipelago was colonized a single time by a species of Saxifraga originating from the Eurosiberian region. On the other hand, the molecular evidence also suggests that populations of S. globulifera from North Africa have been isolated for a long time from populations occurring in the Iberian Peninsula, and that the endemic S. reuteriana has evolved from the Iberian populations of S. globulifera. The Mediterranean Sea has probably been an effective isolating barrier for some plant groups that occur in Europe and North Africa.

Inferring complex phylogenies using parsimony: an empirical approach using three large DNA data sets for angiosperms.

To explore the feasibility of parsimony analysis for large data sets, we conducted heuristic parsimony searches and bootstrap analyses on separate and combined DNA data sets for 190 angiosperms and three outgroups. Separate data sets of 18S rDNA (1,855 bp), rbcL (1,428 bp), and atpB (1,450 bp) sequences were combined into a single matrix 4,733 bp in length. Analyses of the combined data set show great improvements in computer run times compared to those of the separate data sets and of the data sets combined in pairs. Six searches of the 18S rDNA + rbcL + atpB data set were conducted; in all cases TBR branch swapping was completed, generally within a few days. In contrast, TBR branch swapping was not completed for any of the three separate data sets, or for the pairwise combined data sets. These results illustrate that it is possible to conduct a thorough search of tree space with large data sets, given sufficient signal. In this case, and probably most others, sufficient signal for a large number of taxa can only be obtained by combining data sets. The combined data sets also have higher internal support for clades than the separate data sets, and more clades receive bootstrap support of > or = 50% in the combined analysis than in analyses of the separate data sets. These data suggest that one solution to the computational and analytical dilemmas posed by large data sets is the addition of nucleotides, as well as taxa.