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Transformation of Nematodes With the Helios Gene Gun

DNA transfer rates of up to 1% were observed using this selection method, which became considerably enriched during following generations after further transfer (F3/F4: 20%; F5/F6 up to 90%), proving their genetic fixation.

Besides marker gene rol-6, we have employed two different reporter genes, a mutant form of GFP (green fluorescent protein), which was originally cloned from the jellyfish Aequorea victoria (Prasher et al., 1992; Cody et al., 1993; Chalfie et al., 1994) with enhanced fluorescence (Siemering et al., 1996), and -galactosidase from Escherichia coli. Expression was visualized by standard fluorescence microscopy (GFP; see Figure 4) or by histochemical staining (lacZ).

In both cases we have observed marker gene expression under transcriptional control of the SV40 early promoter (Figure 3 A and B), or of the C. elegans actin-1 core promoter after Helios gene gun mediated delivery of Au/DNA particles into the nematode. The lacZ expression of constructs containing the actin-1 core promoter of C. elegans (Files et al., 1983; Krause et al., 1989; Stone and Shaw, 1993) in DNA transfected L. sigmodontis will even be analyzed histochemically.


Discussion
The Biolistic DNA-transfection procedure, because of its simple mode of handling, has an important potential in various innovative molecular biology applications, such as DNA vaccination, protein targeting, and basic research applications such as gene regulation studies. The new Biolistic technology should therefore be able to substitute for other applications in this field. Also, the prepared DNA/Au particles can be stored for rather long periods, at least up to 8 months, which makes repeti
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