Nucleic Acids Research, Vol 24, Issue 11 2166-2175, Copyright © 1996 by Oxford University Press
KG Belanger, C Mirzayan, HE Kreuzer, BM Alberts and KN Kreuzer
The rolling circle DNA replication structures generated by the in vitro
phage T4 replication system were analyzed using two-dimensional agarose
gels. Replication structures were generated in the presence or absence of
T4 primase (gp61), permitting the analysis of replication forks with either
duplex or single-stranded tails. A characteristic arc shape was visualized
when forks with single-stranded tails were cleaved by a restriction enzyme
with the help of an oligonucleotide that anneals to restriction sites in
the single-stranded tail. After calibrating the gel system with this
well-studied rolling circle replication reaction, we then analyzed the in
vivo replication directed by a T4 replication origin cloned within a
plasmid. DNA samples were generated from infections with either wild-type
or primase-deletion mutant phage. The only replicative arc that could be
detected in the wild-type sample corresponded to duplex Y forms, consistent
with very efficient lagging strand synthesis. Surprisingly, we obtained
evidence for both duplex and single-stranded DNA tails in the samples from
the primase-deficient infection. We conclude that a relatively inefficient
mechanism primes lagging strand DNA synthesis in vivo when gp61 is absent.
ARTICLES
Two-dimensional gel analysis of rolling circle replication in the presence and absence of bacteriophage T4 primase
Department of Microbiology, Duke University Medical Center, Durham, NC 27710, USA.
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  S. K. Leung and J. T. Y. Wong The replication of plastid minicircles involves rolling circle intermediates Nucleic Acids Res., April 1, 2009; 37(6): 1991 - 2002. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. K. Kuo, J. D. Griffith, and K. N. Kreuzer 5-Azacytidine Induced Methyltransferase-DNA Adducts Block DNA Replication In vivo Cancer Res., September 1, 2007; 67(17): 8248 - 8254. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Wanrooij, S. Goffart, J. L.O. Pohjoismaki, T. Yasukawa, and J. N. Spelbrink Expression of catalytic mutants of the mtDNA helicase Twinkle and polymerase POLG causes distinct replication stalling phenotypes Nucleic Acids Res., May 11, 2007; 35(10): 3238 - 3251. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Cohen, N. Agmon, K. Yacobi, M. Mislovati, and D. Segal Evidence for rolling circle replication of tandem genes in Drosophila Nucleic Acids Res., August 9, 2005; 33(14): 4519 - 4526. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 K. C. Dudas and K. N. Kreuzer Bacteriophage T4 Helicase Loader Protein gp59 Functions as Gatekeeper in Origin-dependent Replication in Vivo J. Biol. Chem., June 3, 2005; 280(22): 21561 - 21569. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Bowmaker, M. Y. Yang, T. Yasukawa, A. Reyes, H. T. Jacobs, J. A. Huberman, and I. J. Holt Mammalian Mitochondrial DNA Replicates Bidirectionally from an Initiation Zone J. Biol. Chem., December 19, 2003; 278(51): 50961 - 50969. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. S. Miller, E. Kutter, G. Mosig, F. Arisaka, T. Kunisawa, and W. Ruger Bacteriophage T4 Genome Microbiol. Mol. Biol. Rev., March 1, 2003; 67(1): 86 - 156. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 G. Hong and K. N. Kreuzer An Antitumor Drug-Induced Topoisomerase Cleavage Complex Blocks a Bacteriophage T4 Replication Fork In Vivo Mol. Cell. Biol., January 15, 2000; 20(2): 594 - 603. [Abstract] [Full Text]
|
|
|
|
 |
|
 R. Vaiskunaite, A. Miller, L. Davenport, and G. Mosig Two New Early Bacteriophage T4 Genes, repEA and repEB, That Are Important for DNA Replication Initiated from Origin E J. Bacteriol., November 15, 1999; 181(22): 7115 - 7125. [Abstract] [Full Text]
|
|
|
|
 |
|
 D. M. MacAlpine, P. S. Perlman, and R. A. Butow The high mobility group protein Abf2p influences the level of yeast mitochondrial DNA recombination intermediates in vivo PNAS, June 9, 1998; 95(12): 6739 - 6743. [Abstract] [Full Text] [PDF]
|
|