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Nature of the Complementary Strands Synthesized in Vitro upon the Single-Stranded Circular DNA of Bacteriophage øX174 after Ultraviolet Irradiation

Poddar, Ramendra K. and Sinsheimer, Robert L. (1971) Nature of the Complementary Strands Synthesized in Vitro upon the Single-Stranded Circular DNA of Bacteriophage øX174 after Ultraviolet Irradiation. Biophysical Journal, 11 (4). pp. 355-369. ISSN 0006-3495. PMCID PMC1483972. doi:10.1016/S0006-3495(71)86220-3. https://resolver.caltech.edu/CaltechAUTHORS:20160520-141640663

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Abstract

This paper describes experiments intended to decide whether UV lesions in DNA act as absolute blocks to chain elongation by the Escherichia coli DNA polymerase or only slow down the polymerization process. Ultraviolet (UV)-irradiated, single-stranded (SS) circular DNA of bacteriophage øX174 was used as template for the polymerase in a reaction mixture in vitro, under conditions allowing synthesis of not more than one complementary strand per template molecule. The mean length of the newly synthesized complementary strands (as determined by velocity sedimentation in alkaline CsCl gradients), as well as the over-all template activity (as measured by deoxyadenosine monophosphate [dAMP] incorporation) was found to decrease with the number of biologically lethal hits sustained by the irradiated templates. With the increase of time or temperature of reaction, the net synthesis of complementary strands increased (as a consequence of increased initiation), but their mean length remained constant. The mean length of synthesized strands was greater than would be expected if all biologically lethal hits were to block the polymerization process. The lethal hits which serve as blocking lesions are inferred to be pyrimidine dimers because it is possible to obtain synthesis of full-length complementary strands if, when heat-denatured, UV-irradiated, double-stranded replicative form (RF II) DNA of bacteriophage øX174 is used as a template, it is pretreated with yeast photoreactivating enzyme (YPRE) in presence of visible light.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1016/S0006-3495(71)86220-3DOIArticle
http://www.sciencedirect.com/science/article/pii/S0006349571862203PublisherArticle
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1483972/PubMed CentralArticle
Additional Information:© 1971 The Biophysical Society. Received for publication 27 October 1970 and in revised form 17 December 1970. The authors are greatly indebted to Dr. L. B. Dumas, Dr. J. K. Setlow, and Mr. M. Eisenberg for generously supplying us with enzyme and DNA preparations, without which this study would not have been possible. We are also thankful to Dr. Dumas for his enthusiastic help in initiating this work. This research was supported in part by grant GM13554 from the U.S. Public Health Service.
Funders:
Funding AgencyGrant Number
U.S. Public Health Service (USPHS)GM13554
Issue or Number:4
PubMed Central ID:PMC1483972
DOI:10.1016/S0006-3495(71)86220-3
Record Number:CaltechAUTHORS:20160520-141640663
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20160520-141640663
Official Citation:Ramendra K. Poddar, Robert L. Sinsheimer, Nature of the Complementary Strands Synthesized in Vitro upon the Single-Stranded Circular DNA of Bacteriophage øX174 after Ultraviolet Irradiation, Biophysical Journal, Volume 11, Issue 4, 1971, Pages 355-369, ISSN 0006-3495, http://dx.doi.org/10.1016/S0006-3495(71)86220-3. (http://www.sciencedirect.com/science/article/pii/S0006349571862203)
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:67218
Collection:CaltechAUTHORS
Deposited By: George Porter
Deposited On:20 May 2016 21:28
Last Modified:11 Nov 2021 00:29

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