Sunflower (Helianthus annuus) leaves contain compounds that reduce nuclear propidium iodide fluorescence

Sunflower leaves have unidentified compounds that interfere with propidium iodide (PI) intercalation and/or fluorescence. Independently prepared pea leaf nuclei show greater PI fluorescence than nuclei from pea leaves simultaneously processed (co-chopped) with sunflower leaves. Differences in fluorescence persist after mixing the PI-stained pea and the co-chopped pea/sunflower samples, i.e. PI staining protects the nuclei from the effects of the inhibitor. The current results are significant to practical flow cytometric determination of plant nuclear DNA content. They show: (1) simultaneous processing of nuclear samples from the target and the standard species is necessary to obtain reliable DNA estimates; (2) a test for the presence of inhibitors should be conducted; and (3) when inhibitors are present caution should be taken in interpreting differences in estimated DNA content. The previously reported environmentally-induced variation in DNA content in sunflower populations is most simply explained by variation in the amount of environmentally-induced inhibitor that interferes with intercalation and/or fluorescence of PI. Intraspecific variation of DNA content for Helianthus annuus needs to be re-evaluated using best practice techniques comparing physiologically uniform tissues that are free of inhibitors. The best estimate for 2C DNA content of H. annuus used in this study is 7.3 pg. (C) 2000 Annals of Botany Company.