Nuclear Transplantation in Amoebae. I. - Journal of Cell Science

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452 Lorch and Danielli—Nuclear Transplantation in Amoebae. I and colour of the cytoplasm by transmitted light. None of these features was found to be of decisive diagnostic value for the particular cultures used in this study. It may be noted at this point that descendants of a culture of A. proteus obtained from Dr. J. A. Dawson of New York in 1950 correspond much more closely to Schaeffer's description of this species than does the Glasgow stock. A very noticeable difference between the two species is the size of the animals: specimens of A. proteus are in general about one-third longer than A. discoides. No volume measurements were made. Size differences may be useful in distinguishing between cultures of the two species but cannot be applied to individuals, as there is considerable variation. It was thought that dividing amoebae of the two species might display some characteristic difference and therefore observations and camera lucida sketches of a considerable number of dividing amoebae were made (Doljanski, 1954). The mode of cytoplasmic division in both species was found to be.as described by Dawson and others (1937) and Chalkley and Daniel (1933) for A. proteus. Again no decisive difference between the two species was found. Thus it is seen that no means of distinguishing between a single Amoeba proteus and a single A. discoides was found from an examination of the locomotion of the animal or the appearance of the cytoplasm, but flourishing cultures could be distinguished readily by sorting outline drawings of random samples. The possibility of finding characteristic differences between the nuclei of the two species was then investigated. In both species the nucleus rolls about freely in the internal cytoplasm and is habitually found in the posterior third of the animal. It is prevented from being carried forward in the stream by the network of fine cytoplasmic strands which exists in the anterior part of the amoeba but which is continually being dissolved at the tail end (Goldacre, 1952). The shape of the nucleus oi A. proteus was found to be basically discoid, but nuclei are often folded or notched so that they can take on various shapes. In young specimens (up to about 6 hours after division) the nucleus is discoid and has a smooth outline. It is easily deformed temporarily by the cytoplasmic streaming. In very old amoebae the nucleus is folded into bizarre shapes. The nucleus of A. discoides is discoid and is neither folded nor notched except in very old or starved specimens. Our observations on the interphase nuclei of the two species agree with those of Schaeffer (1916) and of Hayes on A. discoides (1938). The fact which emerges from a study of the shapes of nuclei is that if a young active amoeba has a notched or folded nucleus, then it is almost certainly A. proteus. But an old amoeba with a folded nucleus, or any amoeba with a smooth discoid nucleus, cannot be readily identified in this way. The size of the nuclei, like the volume of the amoeba, is greater in A. proteus than in A. discoides. Schaeffer (1916) gives the average diameters of the nuclei of two species as 46 by 15 JU. for proteus and 40 by 18 /x'for discoides. He does not state on how many measurements these averages were based, nor the degree of variation.
Lorch and Danielli—Nuclear Transplantation in Amoebae. I 453 Since the nucleus increases its volume between two divisions, it is necessary to measure a number of nuclei in order to get an average value for a particular culture. Measurements of the largest diameter of the nuclei of random samples taken from cultures of the two species were made, and the distribution of diameters plotted. Only mononucleate amoebae were used. The experiments were designed to find out whether the distribution of diameter and the average nuclear diameter are constant for particular clones under different conditions, or vary with the age, state of nutrition, &c. of the culture; whether the average 30 32 34 36 38 4-0 42 44 46 48 50 52 54 56 58 60 62 64 nuclear diameter in ^ FIG. 3. Nuclear diameters of amoebae in mass cultures compared with those in rapidly growing cultures. Solid line: A. proteus in mass cultures (cultures P2—5, table 2). Broken line: A. proteus in rapidly growing cultures (serial numbers P13-15, table 1). nuclear diameter reflects the size of the parent nucleus; and whether there is a significant difference between cultures of A. proteus and A. discoides. Tables 1-4 (see end of paper) and figs. 3 and 4 illustrate the results of this study. The cultures are grouped as 'rapidly growing', i.e. 'young' clones (tables 1 and 3) and 'mass cultures' (tables 2 and 4). The 'young' clones were set up from a single amoeba not more than 3 weeks before the measurement of the nuclei and contain between 50 and 120 amoebae. The 'mass cultures' contain over 1,000 amoebae in a 10 in. culture dish. It is seen that there is no great difference between the average nuclear diameters of recently established and older clones. The former tend to contain more amoebae with small nuclei: the rate of division is often higher in 'young' clones. Fig. 3 illustrates this difference for A. proteus: the graph for the rapidly growing 'young' clones is somewhat displaced to the left. In A. discoides the displacement of the curve for 'young' clones was less marked. Thus age of a culture by itself has no great influence on the distribution of nuclear diameter nor on the average, and we felt justified in representing all the mass cultures together (fig. 4) in order to show the average pattern of distribution of nuclear diameters in A. proteus and A. discoides.
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