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As a summary, a typical DNA rolling circle replication has five steps: [2] Circular dsDNA will be "nicked". The 3' end is elongated using "unnicked" DNA as leading strand (template); 5' end is displaced. Displaced DNA is a lagging strand and is made double stranded via a series of Okazaki fragments. Replication of both "unnicked" and displaced ...
In the single stranded DNA viruses—a group that includes the circoviruses, the geminiviruses, the parvoviruses and others—and also the many phages and plasmids that use the rolling circle replication (RCR) mechanism, the RCR endonuclease creates a nick in the genome strand (single stranded viruses) or one of the DNA strands (plasmids).
This is accomplished by rolling circle replication with the Phi 29 DNA polymerase which binds and replicates the DNA template. The newly synthesized strand is released from the circular template, resulting in a long single-stranded DNA comprising several head-to-tail copies of the circular template. [ 10 ]
The virus replicates through an dsDNA intermediate initiated by the Rep protein. Two major genes are transcribed from open reading frame (ORF) 1 and 2. ORF1 encodes Rep and Rep' for initiation of rolling-circle replication; ORF2 encodes Cap, the only structural and most immunogenic protein forming the viral capsid. [15]
The observed DNA replication intermediates included circular and branched circular concatemeric structures that likely arose by rolling circle replication. When assembling concatemers from synthetic oligonucleotides, increasing salt concentration to 200 mM was found to be a major optimizing factor due to its ability to enhance ionic strength ...
The polymerase is a monomeric protein with two distinct functional domains. Site-directed mutagenesis experiments support the proposition that this protein displays a structural and functional similarity to the Klenow fragment of the Escherichia coli Polymerase I enzyme; [3] it comprises a C-terminal polymerase domain and a spatially separated N-terminal domain with a 3'-5' exonuclease activity.
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Like other large dsDNA viruses, P22 first builds a protein "procapsid" structure and then packages it with the DNA chromosome. [3] P22 procapsid is assembled by a well-studied protein. [3] About 250 molecules of scaffolding protein are present in the procapsid during assembly, but during DNA packaging, the scaffolding protein is released. [3]