Rapid methods to extract DNA and RNA from Cryptococcus neoformans
Rapid methods to extract DNA and RNA from Cryptococcus neoformans
复制标题
DOI：
10.1016/s1567-1356(01)00030-7
复制
发表时间：
2001-12-01
影响因子：
3.2
通讯作者：
Cardinali, Gianluigi
中科院分区：
文献类型：
--
作者：
Bolano, Alessandro;Stinchi, Silvia;Cardinali, Gianluigi
从致病性酵母新型隐球菌中提取核酸通常受到厚且耐药的荚膜的阻碍，所述荚膜占整个细胞体积的至少70%。本文介绍了一种基于机械破碎法提取C.新形动物和其他被包囊的物种。所提出的DNA提取系统包括通过短时间尿素处理和珠粒撞击的胶囊松弛。这两个步骤允许一致的提取，甚至从耐其他程序的菌株。所提出的方法获得的DNA的产量和质量高于那些获得的两个较早描述的方法。该方案可以扩展到每个酵母物种，特别是那些难以处理胶囊的存在。在初步珠击处理后，使用原始裂解基质和FastPrep系统（Bio101）完成RNA纯化。从15 ml过夜培养物（10（9）个细胞）中的产量范围约为1 mg RNA，RNA似乎未降解，使其适合于分子操作。(c)2001年，欧洲微生物学会联合会。由Elsevier Science B.V.出版，版权所有。
Extraction of nucleic acids from the pathogenic yeast Cryptococcus neoformans is normally hampered by a thick and resistant capsule, accounting for at least 70% of the whole cellular volume. This paper presents procedures based on mechanical cell breakage to extract DNA and RNA from C. neoformans and other capsulated species. The proposed system for DNA extraction involves capsule relaxation by means of a short urea treatment and bead beating. These two steps allow a consistent extraction even from strains resistant to other procedures. Yield and quality of DNA obtained with the proposed method were higher than those obtained with two earlier described methods. This protocol can be extended to every yeast species and particularly to those difficult to handle for the presence of a capsule. RNA purification is accomplished using an original lysing matrix and the FastPrep System (Bio101) after a preliminary bead beating treatment. Yields range around 1 mg RNA from 15 ml overnight culture (10(9) cells), RNA appears undegraded, making it suitable for molecular manipulations. (c) 2001 Federation of European Microbiological Societies. Published by Elsevier Science B.V. All rights reserved.