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Improved methods for the formation and stabilization of R-loops

Kaback, David B. and Angerer, Lynne M. and Davidson, Norman (1979) Improved methods for the formation and stabilization of R-loops. Nucleic Acids Research, 6 (7). pp. 2499-2517. ISSN 0305-1048. http://resolver.caltech.edu/CaltechAUTHORS:KABnar79

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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.


Item Type:Article
Additional Information:Copyright © 1979 Oxford University Press. Received 23 January 1979. We are indebted to Fiona Gibson, Lynna Hereford and Michael Rosbash for Poly(A)+ RNA; the Paul D. Elder Co. for Trioxsalen; and Peter Chandler, James Casey and Pauline Yen for valuable discussions. DBK was supported by a Damon Runyon-Walter Winchell Cancer Research Foundation Fellowship and an U.S.P.H.S. National Research Service Award. This research was supported by grants GM 10991 and GM 20927 from the U.S.P.H.S. and RR 07003, Biomedical Research Grant.
Record Number:CaltechAUTHORS:KABnar79
Persistent URL:http://resolver.caltech.edu/CaltechAUTHORS:KABnar79
Alternative URL:http://nar.oxfordjournals.org/cgi/content/abstract/6/7/2499
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:1926
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:23 Feb 2006
Last Modified:26 Dec 2012 08:46

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