A reinvestigation of 5' leads to 3' polarity in 40S ribosomal RNA precursor of Xenopus laevis

Abstract

The 5' leads to 3' polarity of the 40S precursor rRNA molecule relative to the location of the 18S and 28S RNA regions in the precursor has been reinvestigated. Fragments of rDNA derived by the restriction endonuclease EcoRI and cloned in E. coli were partially digested with the exonuclease induced by bacteriophage lambda and with exonuclease III from E. coli. The resulting rDNA fragments with single-stranded tails were hybridized separately with 18S and 28S rRNA, and the formation of the hybrid was monitored by determination of radioactivity and by electron microscopy. Since the location of the EcoRI sites in rDNA is known, and the specificity of the two exonucleases for 5' and 3' ends of DNA strands has been established, the hybridization of the different partially digested rDNA fragments with either 18S or 28S rRNA could be interpreted in terms of polarity of the coding strand of rDNA, and consequently of the RNA (see models in Figure 1). The results supports the following model for the rRNA precursor moleculre: 5' end-transcribed space-18S gene-transcribed space-28S gene-3' end.