Nuclear DNA content and genome size are key factors in biology and biodiversity, and have important implications in modern molecular genetic studies. Diesel tree (Copaifera officinalis L.) and switchgrass (Panicum virgatum L. cv. Alamo) are attractive sources for biofuels production, including biodiesel and cellulosic ethanol. On the other hand, horseweed (Conyza canadensis L.) is one of the most agriculturally problematic herbicide-resistant weeds worldwide. The nuclear DNA contents (expressed as 1C values) of these economically significant plants were estimated by a fast and valid method of laser flow cytometry. Intact nuclei were isolated from young leaves or roots, stained with propidium iodide, a fluorescent DNA-staining dye, and then analyzed by a flow cytometer simultaneously with nuclei from reference standard plants of known C-values. The 1C-value of the nuclear DNA content of C. officinalis was ca. (1,161 Mbp = 1.19 pg DNA). It is approximately similar to that of oilseed rape and soybean, and is more than eight times the genome size of Arabidopsis thaliana (L.) Heynh. ecotype Columbia. The 1C DNA content of switchgrass (P. virgatum) cv. ‘Alamo’ tetraploid ecotype was estimated at 1,445 Mbp (= 1.48 pg DNA), which is more than ten times the genome size of A. thaliana. Finally, the DNA content of the diploid horseweed (C. canadensis) was ca. (378 Mbp = 0.39 pg DNA), which is approximately 2.8 times the genome size of A. thaliana, and smaller than the reference genomes of rice and poplar. These data will be useful for molecular and genomic approaches, such as genome sequencing projects and construction of genomic libraries.