Robustly Cycling Cell-Free Extracts in Teflon Chambers.

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.