We have developed a new primer design method based on the QuickChange site-directed mutagenesis protocol, which significantly improves the PCR amplification efficiency. This design method minimizes primer dimerization and ensures the priority of primer-template annealing over primer self-pairing during the PCR. Several different multiple mutations (up to 7 bases) were successfully performed wit...

The yeast Ty1 LTR retrotransposon replicates by reverse transcription and integration; the process shows many similarities to the retroviral life cycle. However, we show that plus strand strong-stop DNA transfer in yeast Ty1 elements differs from the analogous retroviral process. By analysis of the native structure of the Ty1 primer binding site and by a series of manipulations of this region a...

The Tf1 retrotransposon represents a group of long terminal repeat retroelements that use an RNA self-primer for initiating reverse transcription while synthesizing the minus-sense DNA strand. Tf1 reverse transcriptase (RT) was found earlier to generate the self-primer in vitro. Here, we show that this RT can remove from the synthesized cDNA the entire self-primer as well as the complete polypu...

A simple and accurate genomic primer extension method has been developed to detect ultraviolet footprinting patterns of regulatory protein-DNA interactions in mammalian genomic DNA. The technique can also detect footprinting or sequencing patterns introduced into genomic DNA by other methods. Purified genomic DNA, containing either damaged bases or strand breaks introduced by footprinting or se...

BACKGROUND Complete mitochondrial (mt) genome sequencing is becoming increasingly common for phylogenetic reconstruction and as a model for genome evolution. For long template sequencing, i.e., like the entire mtDNA, it is essential to design primers for Polymerase Chain Reaction (PCR) amplicons which are partly overlapping each other. The presented chromosome walking strategy provides the over...

Background: Torque teno virus (TTV) was the first human Anelloviridae detected in a Japanese patient with an unknown type of hepatitis in 1997. TTV is by far the first known single-stranded circular DNA virus infecting human. In spite of its widespread nature in human population, its pathogenesis is still unclear. In addition, information regarding TTV infection in Iranian population is limited...

Because of its importance for the origin of life, the nonenzymatic copying of RNA templates has been the subject of intense study for several decades. Previous characterizations of template-directed primer extension using 5'-phosphoryl-2-methylimidazole-activated nucleotides (2-MeImpNs) as substrates have assumed a classical in-line nucleophilic substitution mechanism, in which the 3'-hydroxyl ...

In addition to their capacity for template-directed 5' --> 3' DNA synthesis at the polymerase (pol) site, DNA polymerases have a separate 3' --> 5' exonuclease (exo) editing activity that is involved in assuring the fidelity of DNA replication. Upon misincorporation of an incorrect nucleotide residue, the 3' terminus of the primer strand at the primer-template (P/T) junction is preferentially t...

We propose and investigate a formal language operation inspired by the naturally occurring phenomenon of DNA primer extension by a DNAtemplate-directed DNA Polymerase enzyme. Given two DNA strings u and v, where the shorter string v (called primer) is Watson-Crick complementary and can thus bind to a substring of the longer string u (called template) the result of the primer extension is a DNA ...