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The importance of DNA synthesis during the early development of the sea urchin egg is discussed, as compared with the constancy of many other cellular constituents. The amount of enzymes which deaminate dCMP and MedCMP has been determined in different stages of development of the sea urchin embryo. A striking decrease in the two deaminases was observed, see Fig. 6. The following hypotheses have been discussed. (1) A cycle of conversion of the pyrimidine deoxynucleotides may exist in multiplying cells. In this way an appropriate assortment of pyrimidine deoxynucleotides is produced for DNA synthesis. (2) The deamination of dCMP to dUMP may have a pacemaker function for the DNA synthesis.

Autoradiographic experiments undertaken with 3H-thymidine, 3H-uridine and 3H-cytidine in order to follow nucleic acid metabolism in developing sea urchin eggs have shown that, during the first steps (cleavage) of development, a mechanism of rapid DNA synthesis must exist in the egg. The metabolically active RNA, strictly restricted to the nucleus, that we have demonstrated in these young stages could be a “messenger RNA”.

Separation of nucleic acid derivatives by thin-layer electrophoresis on a microcrystalline cellulose, Avicel, is reported. The present paper describes: 1. 1. Separation of the four 2′, 3′-ribonucleotides from an RNA alkaline hydrolysate. 2. 2. Separation of the four deoxynucleotides and the four deoxynucleosides. 3. 3. Separation of deoxynucleoside mono-, di- and triphosphates. 4. 4. Separation of ten dinucleotides from a deoxyribonuclease digest. 5. 5. Two-dimensional separation of 13 bases occurring in nucleic acids.

1. The amount of deCMP-deaminase (deoxycytidylate aminohydrolase) per embryo has been determined in different stages of development of Rana esculenta. An increase in the content of enzyme per embryo was observed during development. The enzymic content increases approximately twice during the period from cleavage to the end of gastrulation, is relatively constant during neurulation until after hatching, and finally at stage 27 increases to a level twice that of the previous stage. 2. Experiments were made with two batches of eggs at two temperatures (15° and 18°). The development is more rapid at the higher temperature and the levels of activity are generally higher at the higher temperature, but there were no significant changes in the enzymic pattern. 3. Comparisons of gynogenetic haploids with normal diploids were made. No significant difference was observed between haploid and diploid patterns of activity.

Quantitative determinations of the total RNA and of the RNA of the subcellular fractions of the developing sea urchin embryo are reported. By differential centrifugation a nuclear fraction, a mitochondrial fraction, two microsomal fractions and a supernatant fraction have been prepared. It has been found that in the early embryonic development the total RNA of the embryo does not change, whereas an increase of the nuclear RNA and an equivalent decrease of the cytoplasmic RNA occurs. Pulse experiments with 32P orthophosphate at several stages of the early embryonic development are reported. At all stages of development the RNA of the subcellular fractions incorporate the 32P orthophosphate in a way peculiar to each fraction.

Thin-layer chromatographic separations on a new layer, microcrystalline cellulose “Avicel”, and on DEAE-cellulose are presented. They include: 1. 1. R F values of 15 bases and methylated analogues of the bases in six different solvents, of 11 nucleosides in 5 different solvents, and of 8 nucleotides in two solvents. 2. 2. Separation of mixtures of deoxynucleoside and ribonucleoside, mono-, di-, and tri-phosphates of the adenine, guanine, uracil, and cytosine series. 3. 3. Separation of mixtures of the four ribonucleoside 2′,3′-monophosphates, of the four ribonucleosides 5′-monophosphates, and of the four deoxyribonucleoside 5′-monophosphates. The advantages of microcrystalline cellulose over other materials for thin-layer chromatography are discussed.