Abstract
This chapter describes a system that fulfills many of the requirements for the study of gene control in vitro . The system makes possible the study of the primary transcription event separate from later maturation and transport events in RNA synthesis. While the results in the chapter are described for myeloma cells, lines derived from the MPC-11 tumor, the preliminary results with chick embryos, and those of others with HeLa cells and other myeloma cells suggest that these methods may be generally applicable. There are many examples of systems of isolated nuclei where the RNA product is small. Moreover, the isolated nuclei synthesize a very small product, but the nuclear subfraction they isolate (nucleoli) synthesizes very large RNA. This chapter illustrates that crude homogenates and crude nuclei have minimal RNA synthetic activity but activity becomes evident on further purification. Thus, in each system careful evaluation of conditions for optimal activity is required and they may well vary extensively.
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