The family Hyacinthaceae constitutes about 900 species of bulbous geophytes usually characterized by high mucilage content. Taxonomic delimitation of Hyacinthaceae has been controversial since the time of Linnaeus due to the absence of reliable discriminating characters. Pattern of genome size variation can thus be considered as an added character to aid intergeneric and intrageneric relationship of the group. However, reports on genome size estimation by flow cytometric analysis of these plants are rare due to the presence of mucilage, which causes problem with nuclei sample preparation. To overcome this problem five reported nuclei isolation buffers were tested in Drimia indica of which Galbraith's buffer gave comparatively better results and was further modified by increasing pH, detergent concentration, and replacing sodium citrate by citric acid. The modified buffer enabled better sample preparation with increased yield, lesser debris, and improved DNA peak CV. The standardized buffer was used to estimate the 2C values of Drimia indica, Drimia nagarjunae, Drimia wightii, Drimia coromandeliana, and Ledebouria revoluta for the first time by flow cytometric analysis. This study also opens up the scope for further improvement in sample preparation for flow cytometric analysis of mucilaginous plants, which is otherwise problematic due to nuclei clumping and increased viscosity of sample. © 2014 International Society for Advancement of Cytometry.