Worm Breeder's Gazette 8(3): 94
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
We are attempting to develop a transformation system in C. elegans using the cloned sup-7 gene. The sup-7 gene was chosen since its expression will provide a selection of transformed animals among the progeny of injected animals. In our first experiments, animals homozygous for e1107, an amber mutation in tra-3, were microinjected with pJK2 (a 9.8kb plasmid constructed from the original plasmid provided by R. Waterston). Self progeny of homozygous e1107 animals produce only pseudomales. Of 24 animals injected, 2 produced suppressed progeny. Suppression was scored by looking for a perfect hermaphrodite tail under the dissecting scope and confirming this with Normarski. Controls performed during tRNA injections (Kimble et al., 1983) showed that e1107 pseudomales never have either perfect hermaphrodite tails or perfect hermaphrodite gonads. Among the transformed progeny of one injected animal, 3 had normal shaped hermaphrodite gonads producing only sperm and 6 had intersexual gonads producing only sperm and 6 had intersexual gonads producing only sperm. [repeat in original] Thus, although suppressed, the animals were sterile. We next injected pJK2 into animals carrying e364, an amber allele of dpy-13. The injected strain also carried e678 so that suppression of the Dpy phenotype might generate Lon animals to facilitate the screen for suppression. Uninjected e364 e678 animals produced 0/105 Lon animals in a controlled screen. After injection, no Lon worms were found in F1 or F2 progeny, but Lon worms were found in the F3. In one injection, Lon animals were seen through F6, but F7 and F8 progeny were all Dpy. Lon animals appeared sick, but produced small broods containing mostly Dpy and a few Lon worms. Dot blots of DNA isolated from the F1-F3 progeny of the injected worms showed that a yeast sequence present on the plasmid as a molecular marker was also present in each of the populations with Lon worms. (However, this sequence was also found in some populations with no Lon worms.) Uncut DNA isolated from populations with Lons was used to transform bacteria selecting for ampicillin resistance. These bacterial transformants contained the same plasmid that was injected, as judged by restriction digestion. These results show that injected DNA is inherited through several generations and that the injected sup-7 gene is expressed. We are currently cloning random nematode DNA fragments into a plasmid containing the sup-7 gene in an attempt to stabilize the plasmid and thereby reduce the deleterious effects of sup-7 expression.