Worm Breeder's Gazette 10(3): 161
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.
Individual embryos in the pronuclear stage have been attached to a polylysine-coated microscope slide and centrifuged in a predetermined orientation at 16.000g. For this the slide is fixed in the slot of a plastic container which fits exactly into the metal bucket of a swing- out rotor. Under these conditions the eggs reliably stick to the glass surface and slides do never break. At a starting temperature of 80 C the eggs are centrifuged for 15 min. During this time they usually cleave once. Longer centrifugation at lower temperatures can suppress cleavage but obviously not replication of centrioles because frequently tetrapolar cleavage takes place after transfer to room temperature. Using the parameters stated above, eggs were centrifuged in three different orientations: a. from posterior to anterior, b. from anterior to posterior, c. perpendicular to the centrifugal axis. In all cases three distinct cytoplasmic layers formed during centrifugation: at the centrifugal pole a densely packed area of yolk granules, a granula-free area in the middle and lipid droplets concentrated at the centrifugal pole. Neither yolk granules nor lipid droplets appear to be essential for cleavage. Those cells which had received essentially clear cytoplasm nevertheless were found to cleave with the same cell cycle periods as others and appear to produce a comparable number of cells. Centrifugation from the posterior to the anterior pole of the egg results in an unequal cleavage, AB always being too large relative to P1. Consequently, abnormal cell cycle rhythms are expressed such that in AB descendants they are accelerated and P1 descendants they are retarded, leading to an altered sequence of cleaving cell groups and early pattern defects. After centrifugation from the anterior to the posterior pole, different types of 2-cell stages result. These include an enlarged posterior cell with faster cell cycle periods than the smaller anterior cell or, alternatively, the generation of two cells of similar size and behavior. None of them expresses germline quality. After a-p centrifugation often a cytoplast containing most of the yolk granules forms at the posterior pole which does not participate in cleavage. Consequently, monsters with several hundred cells develop which may express visible signs of differentiation (e.g gut birefringency and muscle twitching). Centrifugation perpendicular to the long axis of the egg gave results similar to centrifugation from anterior to posterior. Attempts to obtain two P1-like cells (like in Boveri's experiments with Ascaris eggs ) have been unsuccessful so far. Analysis of P granule distribution and of development after pulsed application of cytoskeletal inhibitors prior to centrifugation are underway.