Pilcher Karen E, Fey Petra, Gaudet Pascale, Kowal Anthony S, Chisholm Rex L
dictyBase, Center for Genetic Medicine, Northwestern University, 676 North Saint Clair Street Suite 1260, Chicago, Illinois 60611, USA.
Nat Protoc. 2007;2(6):1325-8. doi: 10.1038/nprot.2007.180.
In this protocol, we present a standard method for extracting DNA from cells of the social amoeba Dictyostelium discoideum. While this procedure is similar to other phenol:chloroform-based purification methods, it is modified to account for the high level of carbohydrate and nucleases found in Dictyostelium cells. Genomic DNA can be isolated from wild-type and genetically modified cells using the described protocol, allowing molecular genetic analyses to be performed. Following cell lysis, nucleic acid extraction, and precipitation, the isolated DNA is suitable for digestion by restriction enzymes, amplification by PCR and Southern blotting. This procedure takes approximately 3 h to complete.
在本实验方案中，我们介绍了一种从社会变形虫盘基网柄菌的细胞中提取DNA的标准方法。虽然该程序与其他基于酚：氯仿的纯化方法相似，但进行了修改以适应盘基网柄菌细胞中发现的高水平碳水化合物和核酸酶。使用所述方案可以从野生型和转基因细胞中分离基因组DNA，从而进行分子遗传学分析。经过细胞裂解、核酸提取和沉淀后，分离出的DNA适用于限制性内切酶消化、PCR扩增和Southern印迹分析。此过程大约需要3小时完成。
