Enrichment and Characterization of the DNA Coding for Vitellogenin in Xenopus laevis

Abstract

Purified vitellogenin mRNA of Xenopus laevis was incubated with mechanically sheared DNA in high concentrations of formamide and the resulting R‐loops (i.e. RNA · DNA hybrid fragments) separated from the bulk DNA by caesium chloride buoyant density centrifugation. Hybridization with ¹²⁵I‐labeled vitellogenin mRNA revealed a 15–30‐fold enrichment of the DNA coding for vitellogenin. Restriction analysis of the R‐loop‐enriched DNA demonstrated that all known endonuclease HindIII fragments coding for vitellogenin of unfractionated Xenopus DNA were also present in the enriched material, including the specific fragments for the oligo(A)‐containing segment of the RNA. Comparison of these restriction data with the structure found in cloned vitellogenin cDNA, indicates the presence of at least one intervening sequence in the genomic DNA coding for vitellogenin.