The arrangement of length heterogeneity in repeating units of amplified and chromosomal ribosomal DNA from Xenopus laevis

Abstract

Non-transcribed spacer regions of Xenopus laevis ribosomal DNA have been found which vary in length between 1.8 × 10 6 and 5.5 × 10 6 daltons. Length variation of rDNA † † Abbreviations used: rDNA, ribosomal DNA; rRNA, ribosomal RNA; SET, 0.15 m-NaCl, 0.05 m-Tris · HCl (pH 7.8), 0.01 m-EDTA; SDS, sodium dodecyl sulfate; cRNA, complementary RNA, transcribed in vitro with Escherichia coli RNA polymerase. repeats exists within a single nucleolar organizer. Amplified rDNA contains repeats of the same size classes but often in different abundance than the chromosomal rDNA of the same animal. If a certain repeat length is preferred during amplification in an individual, it is also preferred in siblings with the same chromosomal rDNA composition. Thus, preference for a size class in amplification is inherited. Some animals selectively amplify repeat lengths which are rarely found in their chromosomal rDNA; others amplify their most abundant size class. The intramolecular arrangement of length variability was analyzed by the electron microscopy of heteroduplex molecules. Long single strands from two separate preparations of amplified and chromosomal rDNA each were reannealed with an homogeneous cloned spacer-containing rDNA fragment (CD30), and the size of adjacent heteroduplex regions was determined. The arrangement of length heterogeneity is very different in the two types of rDNA. Most, if not all, tandem repeats along a single molecule of amplified rDNA are equal in length. This observation supports a rolling circle mechanism for amplification. In contrast, between 50% and 68% of adjacent repeats in a given molecule of chromosomal rDNA differ in length. For one of the chromosomal rDNA preparations analyzed, the frequency of non-identical nearest-neighbors is compatible with random scrambling of repeats of different lengths. This result bears on the mechanism by which tandem genes evolve. It rules out sudden correction mechanisms of tandem genes such as the “master-slave” or certain “expansion-contraction” models, which predict that tandem genes will be identical.