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The manuscript entitled ?dynamics of genome size and its co-relation to chromosome number of P. pinnata? shows the optimisation of nuclear isolation buffer exclusively for P. pinnata. Isolation of nuclei from the tender leaves of P. pinnata was really a great challenge as the nuclei was disintegrated in detergents apart from IGEPAL CA- 630 (NP-40). When isolation of nuclei was successfully established, the external standardization was preliminary followed to find the genome size of the P. pinnata. Recent techniques of flow cytometric protocol paved the way to internal standardization. Preliminary experiments of P. pinnata in both internal and external standardization show us there was the inhibition of the P. pinnata cytosol on the mean fluorescence of the standard nuclei. So using the current established protocols the inhibition was proved. The effect of different volume combinations on the nuclear DNA content estimated in pseudo-internal and internal standardization was evaluated. The deviation from the proportional relationship of mean P. pinnata fluorescence and mean standard nuclei fluorescence motivated us to add antioxidants (PVP and 2-mercaptoethanol). Although their addition could not able to alleviate the inhibitory effect of P. pinnata cytosol, their addition reduced the inhibition to the minimum (least percentage shift of the mean standard nuclei fluorescence due to internal standardization and the best fit curve of the antioxidant- nuclear isolation buffer combinations). So the originality of the manuscript lies in the least quantitative shift of the standard nuclei fluorescence and use of the best fit curve of different standardization methods and different antioxidant- nuclear isolation buffer combinations to alleviate the P .pinnata cytosol effect paving the way to accurate estimation of the nuclear DNA content. The authors believe that the paper would be a trend setter in the plant DNA content analysis as there is an urgent attention of estimation of nuclear DNA content of 97 % of the angiosperms.
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