Abstract
Although genetic diseases are usually described in terms of enzyme function and structure, further understanding of genetic mechanisms in the human will also depend upon the ability to physically isolate and characterize specific human genes. We have used thermal fractionations, molecular hybridization and sequential equilibrium density gradients to purify the human genes of 28S and 18S RNA - the nucleic acid components of ribosomes. The genetic sequence was isolated as a rRNA/DNA hybrid which was homogenous as demonstrated by a sharp melting transition at 80° in 0.1 × SSC. Linkage of 28S RNA and 18S RNA genes on the same isolated sequence was demonstrated by isolation of 28S RNA/DNA hybrids, followed by 18S hybridization with that molecule. The specific gravity of the hybrid (1.745) indicates that 30% of the DNA sequence is complimentary to 28 & 18S RNA. The remainder is probably accounted for by transcribed precusor molecules and nontranscribed “spacer” regions. The chromosomal localization of rDNA was possible by a method in which we added 3H-Uridine to human cells auxotrophic for uridine to achieve rRNA of very high specific activity and purity. In situ hybridization of this rRNA with chromosomes demonstrated that the ribosomal genes were located only on the D and G group chromosomes. The potential for studying fine chromosomal structure by this technique is demonstrated by the absence of ribosomal RNA genes on a D/D translocation chromosome.
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Schmickel, R., Knoller, M. DIRECT CHROMOSOMAL LOCALIZATION, ISOLATION, AND CHARACTERIZATION OF A SPECIFIC HUMAN GENE. Pediatr Res 8, 394 (1974). https://doi.org/10.1203/00006450-197404000-00327
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DOI: https://doi.org/10.1203/00006450-197404000-00327