Pilcher Karen E, Gaudet Pascale, Fey Petra, 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):1329-32. doi: 10.1038/nprot.2007.191.
Here we present a protocol for the extraction of RNA from Dictyostelium discoideum. Dictyostelium is a social amoeba that undergoes a basic developmental program, and therefore analysis of RNA levels over a time course is a commonly used technique. This procedure is similar to other guanidine thiocyanate-based methods; however, it has been adjusted because of the large quantities of carbohydrate and nucleases found in Dictyostelium cells. After cell lysis and phenol:chloroform extraction, the resulting high-quality RNA isolated with the described protocol allows the molecular genetic analysis of wild-type and genetically modified cells. The purified RNA can be used for analyses such as northern blotting, RT-PCR and microarrays. This procedure requires approximately 2 h to complete.
在此，我们展示了一种从盘基网柄菌中提取RNA的方法。盘基网柄菌是一种会经历基本发育程序的社会性变形虫，因此在一个时间进程中分析RNA水平是一种常用技术。该方法与其他基于硫氰酸胍的方法类似；然而，由于在盘基网柄菌细胞中发现了大量碳水化合物和核酸酶，所以对其进行了调整。细胞裂解并经苯酚：氯仿抽提后，用所述方法分离得到的高质量RNA可用于野生型和转基因细胞的分子遗传学分析。纯化后的RNA可用于诸如Northern印迹、逆转录PCR和微阵列等分析。此过程大约需要2小时完成。
