Kjs

b nondysgenic crosses

normal normal normal progeny progeny progeny c dysgenic crosses

F, progeny frequency sterile

Fj progeny with many mutations.

FIGURE 21-21 Hybrid dysgenesis.

P-element transposons reside passively in P strains because tfiey exptess a repressor that keeps the transposons silent. When P strains are mated with an M strain lacking such a repressor, the transposons are mobilized within the pole cells, arid often integrate mto genes required fof germ cell formation This explains the high frequency ot sterility m the offspring from such crosses length P-element transposon is 3 kb in length. It contains inverted repeats at the termini thai are essential for excision and insertion. The intervening DNA encodes both a repressor of transposition and a transposase that promotes mobilization. The repressor is expressed in the developing eggs of P strains. As a result, there is no movement of P-elemen!s in embryos derived from females of the P-strain (these contain P-elements). Movement is seen only in embryos derived from eggs produced by M strain females, which lack P-elements.

Recombinant DNA is inserted into defective P-elements that lack the internal genes encoding repressor and transposase. This DNA is injected into posterior regions of early, precellular embryos (as we saw in Chapter IB, this is the region that contains the polar granules). The transposase is injected along with the recombinant P-element vector. As the cleavage nuclei enter posterior regions, they acquire both the polar granules and recombinant P-element DNA together with transposase. The pole cells bud off from the polar plasm and the recombinant P-elements insert into random positions in the pole cells. Different pole cells contain different P-element insertion events. The amount of recombinant P-element DNA and transposase is calibrated so that, on average* a given pole cell receives just a single integrated P-element. The embryos are allowed to develop into adults and then mated with appropriate tester strains.

The recombinant P-element contains a "marker1" gene such as white" and the strain used for the injections is a white mutant. The tester strains are also white . so that any Fz fly that has red eyes must contain a copy of the recombinant P-element. This method of P-element transformation is routinely used to identify regulatory sequences such as those governing eve stripe 2 expression (which we discussed in Chapter 18), In addition, this strategy is used to examine protein coding genes in various genetic backgrounds.

[n summary, Drosophila offers many of the sophisticated tools of classical and molecular genetics that, as we have seen, are available in microbial model systems. One conspicuous exception has been the absence of methods for precise manipulation of the genome by homologous recombination With recombinant DNA, such as in the creation of gene deletions. However, such methods were recently developed, and are now being streamlined for routine use. Ironically, such manipulations are readily available, as we shall see, in the more complicated model system, the mouse, Nevertheless, because of the weallh of genetic tools available in Drosophila and the extensive ground work of knowledge about this organism resulting from decades of investigation,

embryo transformed DNA in gamete genome

P-element embryo transpossse transformed DNA in gamete genome

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