Methods in Drosophila Cell Cycle Biology

The chapter focuses on fluorescent in situ hybridization, immunofluorescence, and primary cultures. As far as fluorescent in situ hybridization is concerned, it includes protocols for the application of this technique to polytene and diploid cells both in squashed and whole-mounted tissues. It deals with immunofluorescence techniques includes protocols for the staining of embryos, third-instar larval brains, and mature oocytes. Finally, a newly developed version of a protocol for primary culture of cells from third-instar larval brains is also included. In situ hybridization to diploid cells provides the only route toward mapping and characterizing the behavior of heterochromatic sequences of Drosophila, which are both underrepresented and aggregated in polytene cells. It includes two protocols for in situ hybridization to diploid cells from squashed and whole-mounted third-instar larval brains. For genome mapping, squashed preparations are by far the best choice because they are easy to make and provide the highest possible resolution. © 1998 Academic Press, Inc.