Comparison of DNA extraction protocols from sour and sweet cherry

The creation of new varieties of fruit crops is impossible without efficient use of genetic resources based on modern methods of molecular biology and genetics, which requires high-quality DNA samples. Extraction of DNA from plant material is a challenging task due to the presence of various inhibitory substances, such as polyphenols and polysaccharides. In this study, we set out to compare four modifications of the original CTAB protocol for DNA extraction from lyophilized leaves of sour cherry (Prunus cerasus) of the Assol, Belyye Zhuravli, and Rusinka varieties and sweet cherry (Prunus avium) of the Podarok Ryazanii, Fatezh, and Chermashnaya varieties. Modifications implying changes in the composition of lysing buffer and washing solution were tested. The original CTAB protocol was used as the control. The extraction was performed in triplicate. DNA concentration and purity were measured using a spectrophotometer; degradation was evaluated by electrophoresis; inhibitors were evaluated using real-time PCR. The conducted study revealed that modification 1 of the CTAB protocol produces DNA samples with the highest degree of purity. An analysis of the A260/280 spectrophotometric index showed values ranging from 1.89 to 1.91 with a mean of 1.87 for modification 1. Modification 2 also yielded high-purity DNA samples, although with the presence of minor RNA amounts in some of them. The A260/230 parameter confirmed the superiority of CTAB modification 1 in obtaining high-purity DNA. The mean value was 2.08, which is comparable to that of the original method. Determination of DNA concentration in aqueous solutions revealed the highest DNA yield when using CTAB modification 1. The concentration ranged from 594.80 to 852.10 μg/mL with a mean of 700.82. Statistical analysis confirmed the pronounced superiority of modification 1 over the control (and other tested modifications) in terms of DNA purity and yield. The results obtained allow us to recommend modification 1 of the CTAB protocol for DNA extraction from plant material of sour and sweet cherry.

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