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Improved methods for the formation and stabilization of R-loops.

D B Kaback·L M Angerer·N Davidson
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Abstract

Improved methods for the formation and stabilization of R-loops for visualization in the electron microscope are presented. The two complementary strands of a duplex DNA are photochemically crosslinked once every 1 to 3 kb using 4, 5', 8 trimethylpsoralen. R-loops are then formed by incubation with RNA in 70% formamide at a temperature above the DNA melting temperature. Finally, the R-loops are stabilized by modifying the free single strand of DNA with glyoxal, thus minimizing the displacement of the hybridized RNA by branch migration. In this manner R-loops can be formed and visualized at a high frequency irrespective of the base composition of the nucleic acid of interest.

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Base SequenceDNAKineticsMicroscopy, ElectronMolecular WeightNucleic Acid ConformationNucleic Acid DenaturationNucleic Acid HybridizationPlasmidsPoly ASaccharomyces cerevisiaeTrioxsalenJournal ArticleResearch Support, U.S. Gov't, P.H.S.

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D B Kaback, L M Angerer, N Davidson. 1979-06-11. Improved methods for the formation and stabilization of R-loops.. https://doi.org/10.1093/nar%2F6.7.2499

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