Uv vis nanodrop

The plant material consisted of 17 Diplotaxis species retrieved from different European genebanks (Figure 1) including the Leibniz-Institut für Pflanzengenetik und Kulturpflanzenforschung, IPK (Gatersleben, Germany), the Royal Botanical Garden Kew (Richmond, United Kingdom), the Universidad Politécnica de Madrid UPM (Madrid, Spain), and the Universidad de Castilla—La Mancha UCLM (Ciudad Real, Spain).
Seeds were sown in pots under climate-grown chamber conditions at the Research Centre for Vegetable and Ornamental Crops (Italy). After 30 days from germination, 100 mg of fresh leaves were collected, followed by nucleic acid isolation using the DNeasy plant mini kit (Qiagen, Hilden, Germany). The DNA purity was estimated by absorbance at 280 and 260 nm, respectively, using a UV-Vis Nanodrop (Thermo Scientific, Wilmington, DE, USA), and the integrity by electrophoresis on a 1.0% agarose gel. Concentration was measured using the Qubit 3 Fluorometer (Thermofisher, Waltham, MA, USA). A volume of 10 μL of extracted DNA, as well as the standards, were diluted in 190 μL of buffer prepared using 1 ul of dsDNA BR Reagent 200 × and 199 μL of dsDNA BR buffer furnished with a Qubit^® dsDNA BR assay kit (Thermofisher, Waltham, MA, USA). DNA was diluted at a working concentration (15 ng/μL) and then stored at −20 °C prior to analysis.

DNA concentration measurements were performed with microvolume nucleic acid quantification (UV–Vis NanoDrop (Thermo Fisher Scientific, Waltham, MA, USA)) using the absorbance measured at 260 nm. To determine the purity of the extracted pDNA, the absorbance of the sample at 280 nm was also measured. A typical ratio of the absorbance at 260 nm and 280 nm (A_260/280) for a pure DNA solution is considered to be between 1.7 and 2.0.