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Recombinant clones specific for ribosomal protein (r-protein) S8 have been isolated from a Xenopus laevis cDNA bank. Sequence analysis shows that they are of two types, derived from two different gene copies originating from gene duplication. The two cDNAs differ in several silent sites and code for the same S8 protein whose complete amino acid sequence has been derived. Sequence comparison of S8 mRNAs with those for other X. laevis r-proteins, has revealed interesting similarities in the 5' and 3' untranslated regions. These could be involved in r-protein synthesis regulation which we have previously shown to occur mainly at post-transcriptional and translational levels.

The interspersion period of repetitive and unique sequences was analyzed by two different methods, electron microscopy and agarose gel electrophoresis, for four Amphibian species with different nuclear DNA content, namely the Anura Xenopus laevis (3 pg DNA per haploid genome) and Bufo bufo (7 pg) and the Urodela Triturus cristatus (23 pg) and Necturus maculosus (52 pg). Within each of the two subclasses it has been found that interspecific differences, in DNA content, due to variations in the amount of repetitive sequences, do not involve variations in length of the interspersed repetitive sequences. They remain about 380 base pairs. Furthermore, the unique sequences length has been found to be shorter in Bufo (760 base pairs) than in Xenopus (1600) and in Necturus (880) than in Triturus (1340). A study of the interspersion period has shown that the great difference in DNA content between Anura and Urodela, which had been previously shown not to have involved changes in the relative amounts of the various sequence classes, does not involve changes in the interspersion period.

Poly-A+ mRNA from Xenopus laevis oocytes, partially enriched for r-protein coding capacity has been used as starting material for preparing a cDNA bank in plasmid pBR322. The clones containing sequences specific for r-proteins have been selected by translation of the complementary mRNAs. Clones for six different r-proteins have been identified and utilized as probes for studying their genomic organization. Two gene copies per haploid genome were found for r-proteins L1, L14, S19, and four-five for protein S1, S8 and L32. Moreover a population polymorphism has been observed for the genomic regions containing sequences for r-protein S1, S8 and L14.

DNA reassociation kinetics were studied, by means of the hydroxyapatite chromatography method, for four species of Amphibians with different nuclear DNA content: Xenopus laevis (3 pg DNA per haploid genome) and Bufo bufo (7 pg) of the Anura subclass and Triturus cristatus (23 pg) and Necturus maculosus (52 pg) of the Urodela subclass. Within each subclass the two species studied were found to have about the same absolute amount of unique DNA. The differences of total nuclear DNA can be accounted for by quantitative variations of the repetitive sequence classes, at least in part due to changes in the number of copies of the various sequences. On the contrary the great difference in nuclear DNA between the two subclasses, Anura and Urodela, involves all sequence classes in parallel; the slowly reassociating fraction appears to be unique in spite of a tenfold difference in absolute amount. The dependence of reassociation kinetics on DNA fragment length for the four species indicates for all of them an interspersed organization of the various sequence classes.

The organization of differently sized genes coding for rRNA in Xenophus laevis has been investigated by partial EcoRI or HindIII digestion. The electrophoretic patterns obtained revealed that most adjacent repeating units are equally sized. This conclusion is in agreement with a previous suggestion that the nucleolar organizer is made up of internally homogeneous blocks of rRNA genes.

Because of the redundancy of ribosomal RNA cistrons and in analogy with a few known special cases, one may postulate that a few "master genes" for rRNA direct the synthesis of multiple copies of themselves per cell cycle. We have shown that in Chinese hamster cells rRNA cistrons duplicate like the bulk of DNA, once in a cell cycle. The time during the S-phase at which rRNA genes duplicate has also been determined by means of RNA-DNA hybridization studies. The time of duplication of rRNA genes occurs between t = 1.5 and t = 3.0 hours from the beginning of the S-phase. [ABSTRACT FROM AUTHOR]