V. H. Sakharova, R. Blume, A. Rabokon, Y. Pirko, Y. Blume
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引用次数: 0
摘要
的目标。本研究的目的是比较不同方法对小茶树标本DNA分离的效率。前女友。,进一步修改这些方法以提高DNA产量,并确定可以提供最佳分离DNA产量的方法。方法。采用DNeasy Plant Mini Kit (QIAgen)和CTAB法对DNA分离方法进行了改进。采用退化引物进行β-微管蛋白内含子长度多态性(TBP)方法的PCR检测。扩增子在聚丙烯酰胺凝胶中分离,然后用硝酸银染色显示。结果。采用CTAB法和DNeasy Plant Mini Kit (QIAgen)的4种修饰方法,成功提取了小叶柏(C. microcarpa)标本叶部和种子的DNA。结论。研究表明,最有效的方法是DNeasy Plant Mini Kit (QIAgen) No. 2。延长细胞裂解期对DNA产率的提高效果最好。我们发现,DNA分离的成功与否与其说是受植物标本室标本年龄的影响,不如说是受植物标本干燥和储存方法的影响。
Comparison of methods of DNA extraction from herbarium specimens of little-pod false flax (Camelina microcarpa Andrz. Ex Dc.)
Aim. The aim of this research was to compare the efficiency of DNA isolation methods from herbarium specimens of Camelina microcarpa Andrz. Ex DC., further modification of these methods to increase DNA yield, and determine the method that would provide the best yield of isolated DNA. Methods. Modifications of the DNA isolation methods using the DNeasy Plant Mini Kit (QIAgen) and the CTAB method were used. PCR was performed using degenerate primers for method of β-tubulin intron length polymorphism (TBP). Amplicons were fractionated in polyacrylamide gel followed by visualization by silver nitrate staining. Results. DNA was successfully extracted from C. microcarpa herbarium specimens sampled with leaf parts and seeds, using the modified by CTAB method, and four modified methods using DNeasy Plant Mini Kit (QIAgen). Conclusions. The study revealed that the most effective method tested was the DNeasy Plant Mini Kit (QIAgen) No. 2. Prolongation of the cell lysis stage had the best effect on the increase of DNA yield. We found that the success of DNA isolation was influenced not so much by the age of the herbarium specimen as by the methods of drying and storing the plants in the collection.