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PMID: 379821 Published · ppublish English Journal Article Research Support, U.S. Gov't, P.H.S.

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

Nucleic acids research ·Vol. 6 ·No. 7 ·1979-06-11 ·Pages 2499-317

Kaback DB, Angerer LM, Davidson N

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.

MeSH Terms
Base Sequence DNA/isolation & purification Kinetics Microscopy, Electron Molecular Weight Nucleic Acid Conformation Nucleic Acid Denaturation Nucleic Acid Hybridization Plasmids Poly A Saccharomyces cerevisiae/analysis Trioxsalen
Chemicals
Poly A DNA Trioxsalen
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Kaback D B
Angerer L M
Davidson N
References (29)
29 references, click to expand
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Article Info
Journal
Nucleic acids research
Abbr.
Nucleic Acids Res
ISSN
0305-1048
DOI
10.1093/nar/6.7.2499
Published
1979-06-11
Pages
2499-317
Language
English
Region
England
NLM ID
0411011
PMCID
PMC327868
Subset
IM
External Links
PubMed source DOI: 10.1093/nar/6.7.2499 PMC Full Text View journal details
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