Chang Jeremy B, Ferrell James E
Department of Chemical and Systems Biology, Stanford University School of Medicine, Stanford, California 94305-5174;
Department of Biochemistry, Stanford University School of Medicine, Stanford, California 94305-5307.
Cold Spring Harb Protoc. 2018 Aug 1;2018(8):pdb.prot097212. doi: 10.1101/pdb.prot097212.
A central advantage of studying is the manipulability of its cell-free extracts, which perform the cell cycle in vitro. However, these extracts are known to be experimentally temperamental and will often complete at most one or two cycles. Over the course of developing systems for imaging cell cycle events in extracts in real time, we unexpectedly found that when standard extracts are placed in Teflon tubes, they cycle extremely robustly; in one series of experiments, over 90% ( = 13) of extracts cycled an average of seven and as many as 14 times. Extracts incubated in other materials, such as glass and polydimethylsiloxane, do not cycle as robustly. Here we present protocols for preparing extracts and imaging them in Teflon tubes. This method extends the usefulness of this powerful model organism.
研究[具体生物名称未给出]的一个核心优势在于其无细胞提取物的可操作性，这些提取物能在体外进行细胞周期。然而，已知这些提取物在实验中不稳定，通常最多只能完成一两个周期。在开发实时成像提取物中细胞周期事件的系统过程中，我们意外地发现，当将标准[具体生物名称未给出]提取物置于特氟龙管中时，它们能极其稳健地循环；在一系列实验中，超过90%（n = 13）的提取物平均循环了七次，多达十四次。在其他材料（如玻璃和聚二甲基硅氧烷）中孵育的提取物则不能如此稳健地循环。在这里，我们展示了制备[具体生物名称未给出]提取物并在特氟龙管中对其成像的方案。这种方法扩展了这种强大模式生物的用途。
