In contrast to many organisms, Drosophila embryos do not integrate injected DNA at an appreciable frequency. For this reason, the generation of germ-line transformants has relied on the utilization of transposable elements to effect the chromosomal integration of injected DNA (1,2). The success of this approach has depended largely on our understanding of the biology of P elements and the syncitial nature of the early Drosophila embryo. The first 13 embryonic divisions following fertilization are nuclear, resulting in the formation of a syncitium. Consequently, if microinjection into the posterior end of the embryo is carried out prior to cellularization, a proportion of the microinjected DNA will be present in the cytoplasm of the pole cells, the progenitor cells of the germ line.
In practice, the DNA to be injected is comprised of two components. The first consists of a helper plasmid containing a defective P element that, although capable of producing the P transposase, which can act in trans on another P transposon, is itself immobile. The second component consists of a transposon construct in which the sequence to be integrated as a transgene is situated between the 31-bp P element inverted terminal repeats along with a suitable marker. The transposase produced by the helper plasmid will act on the inverted repeats of the transposon construct and facilitate the integration of the transposon into essentially random chromosomal sites of the recipient's germ line. Both P element biology and the characteristics of P elementmediated transformation have been reviewed extensively (for example, ref. 3). In this chapter, we will deal primarily with the technical details necessary for obtaining germ-line transformants.
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