Gaudet Pascale, Fey Petra, Chisholm Rex
dictyBase, Center for Genetic Medicine, Northwestern University, Chicago, IL 60611, USA.
CSH Protoc. 2008 Dec 1;2008:pdb.prot5100. doi: 10.1101/pdb.prot5100.
INTRODUCTIONDictyostelium discoideum is a unicellular eukaryote often referred to as a "social ameba" because it can form a multicellular structure when nutrients are depleted from the immediate environment of the cells. In the laboratory, this is accomplished simply by replacing the growth medium with a buffer solution. For best results, it is important to harvest the cells while they are still in exponential growth (1-4 × 10(6) cells/mL). At high cell density, many of the cells in a culture will have initiated development, thus yielding asynchronous development. Dictyostelium cells can be developed on solid media, either on filter paper or on KK2 plates. If only the early stages of development are important (e.g., to study chemotaxis), cells can be developed in suspension. Under these conditions, cells will only progress through the first 6-8 h of development. Addition of cAMP pulses to the starved suspension culture will allow development to progress up to the 12-h stage, corresponding to the beginning of culmination.
引言
盘基网柄菌是一种单细胞真核生物，常被称为“社会性变形虫”，因为当细胞周围环境中的营养物质耗尽时，它能形成多细胞结构。在实验室中，只需用缓冲溶液替换生长培养基就能实现这一点。为获得最佳结果，在细胞仍处于指数生长期（1 - 4×10⁶个细胞/毫升）时收获细胞很重要。在高细胞密度下，培养物中的许多细胞会开始发育，从而导致发育不同步。盘基网柄菌细胞可在固体培养基上发育，如在滤纸上或在KK2平板上。如果仅发育的早期阶段很重要（例如，用于研究趋化性），细胞可在悬浮液中发育。在这些条件下，细胞只会经历发育的前6 - 8小时。向饥饿的悬浮培养物中添加cAMP脉冲将使发育进展到12小时阶段，这对应于聚集开始。
