Chapter 26
FISH on Insect Cells Transfected with Heterologous DNA
THOMAS LIEHR
Introduction
Insect cells transfected with heterologous DNAs (plasmids) can be used
for the assessment of functional properties of genes that have been isolated or subjected to in vitro mutagenesis (Rubin and Spradling 1983,
Snow et al. 1989). An example of a cell system used for such approaches
are Schneider cells (Schneider 1972). These are derived from embryonic
cells of Drosophila melanogaster, can be transfected with special plasmids
(Bunch et al. 1988) and used for heterologous expression. Several, in general, reliable in vitro methods can be performed to control the transfection
efficiency in such cells (i.e. peR, Southem-, Northern- or Western-blot
approaches). Another fast and reliable method to detect transfected
Schneider cells, offering the possibility of assessing the transfection
rate on a single cell level, has recently been described and is based on
a modified FISH technique (Rautenstrauss et al. 1998; see Figure 1)
and described in the following.
Procedure
Preparation of the Schneider cells for FISH
1. Schneider cells transfected for transient expression (e.g. with a derivate
of the plasmid pRmHa-3 (Bunch et al. 1988» and wild type Schneider
cells (without any plasmid, used as negative controls) have to be cultivated according to Schneider (1972).
Thomas Liehr, Institut fur Humangenetik, Kollegiengasse 10, Jena, 007743, Germany
(phone +49-3641-935533; fax +49-3641-935502; e-mail i8Iith@mti-n.mti.uni-jena.de)
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