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Nucleic Acids Research, 1978, Vol. 5, No. 6 2211-2222
© 1978

Articles

Preparation and isolation of covalently closed circular rDNA molecules from DNA of Xenopus laevis

Lino C. Polito and Silvio Spadari*

International Institute of Genetics and Biophysics CNR, Naples **Laboratorio di Genetica Biochimica e Evoluzionistica CNR, Pavia, Italy

Received January 17, 1978. We describe a method leading to the formation of closed circles of rDNA starting from total DNA of Xenopus laevis Linear DNA molecules were digested with exonuclease 3 and self-annealed. Open circles were enriched and covalently closed by the simultaneous use of polynucleotide kinase, DNA polymerase and polynucleotide ligas. Closed circles of rDNA1 were shown to be alkali-resistant, to have higher density than linear molecules in cesium chloride density gradients containing ethydium bromide, and to have the sedimentation constant expected for a single repeat unit of rDNA comprehensive of its spacer.


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