Synthesis of desoxyribonucleic acid during amphibian embryogenesis at different temperatures

Abstract

TITO FIGURES INTRODUCTIONNormal development of amphibian embryos is known to occur within a quite wide temperature interval -around 20°C (cf. Moore, '39). Within this range the morphological development is rather unaffected, although body size may be smaller and gill size larger at high temperatures (Chambers, '08; Doms, '15). This does not imply that the developmental processes are identical at all temperatures. On the contrary it has been possible to show that the supply of energy, and the utilization of energy sources is strongly influenced by temperature (Ldvtrup, '53b ; ' 5 9~) . This in turn influences the synthesis of enzymes in such a way that the maximum amount of enzymes is highest at intermediate temperatures, and decreases as the temperature approaches the limits of the range (Lrzlvtrup, '53c; '55; '59d). Retardation of morphological development may be observed at low temperatures (L@vtrup, '59d). Previous investigations are seen to comprise the following aspects of embryogenesis : Morphological development, energy utilization, and chemical diff ercntiation. I n so far as the content of desoxyribonucleic acid (DNA) may be taken to represent the number of cells, the present investigation of DNA synthesis a t different temperatures represents an analysis of the temperature effect on yet an other embryonic process, viz., cell division. 545 S#REN L~T R U P MATERIAL AND METHODAdult specimens of R a n n pla,tyrrh,ina (= R. temporaria) were collected in nature and allowed to mate in the laboratory. The eggs were placed in Erlenmeyer flasks (50-100 in each), and the flasks were partly left at room temperature (RT; about 20°C), and partly immersed in water baths at 10°C.The DNA content of the embryos was determined on acetone dried powders. The samples from the earliest stages contained 25 eggs or embryos, from the later stages the number was 5-10. The microbiological method developed by Hoff-Jgkgensen ('52; '54) was used for the analyses in the modification suggested by L@vtrup and Roos ( '57). Work on this method which has gone on f o r several years in my laboratory has shown that certain precautions are necessary if reliable results are to be obtained. The most important is probably that the pH and the bu-ffer capacity of the medium must be controlled very carefully. The difference in buffer capacity arising from diluting samples of varying DNA content to different degrees with water suffices to make the results incomparable. It is furthermore our experience that desoxyribonuclease alone cannot digest certain types of DNA sufficiently to make them available for the microorganism. For other types the treatment with this one enzyme may be enough. The additional use of snake venom has been suggested by various authors. We have found that, depending upon the circumstances, snake venom may either increase or decrease the yield, or be without any influence. The details of these investigations m7ill be published in a separate paper (LGvtrup and ROOS, '60). The method used in the present work may not be the ul...