The success of Agrobacterium mediated plant transformation depends to a certain extent on appropriate selection of the A. tumefaciens strain for a particular plant species. Many stages in a plant transformation procedure are prone to bacterial contamination with similar antibiotic resistance that may compromise the identity of the A. tumefaciens strain used, in turn adversely affecting success ...

During the cloning of a transcript of one member of a closely related gene family, the human 7-crystallin gene family (1), we encountered a novel artifact of the polymerase chain reaction: the formation of chimeric cDNA molecules. Our experimental strategy in cloning the human 7E-crystallin transcript was a common one (2): first strand cDNA synthesis on human lens RNA using AMV reverse transcri...

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...

This paper describes the use of real-time PCR for the confirmation of microarray data. Current publication guidelines require that all microarray results are confirmed by an independent gene expression profiling method. Real-time PCR is the method of choice for most researchers but it is not without drawbacks. The first step in confirming array results by real-time PCR is selection of gene-spec...

The Polα/primase complex assembles the short RNA-DNA fragments for priming of lagging and leading strand DNA replication in eukaryotes. As such, the Polα polymerase subunit encounters two types of substrates during primer synthesis: an RNA:DNA helix and a DNA:DNA helix. The engagement of the polymerase subunit with the DNA:DNA helix has been suggested as the of basis for primer termination in e...

In search of a primer set that could be used to study Planctomycetes dynamics in lakes and especially via fingerprinting methods, e.g. denaturing gradient gel electrophoresis (DGGE), three existing specific primer sets, developed for marine and soil systems, have been tested on water samples from four freshwater ecosystems. The first primer set (PLA46F/PLA886R) allowed PCR amplification of Plan...

BACKGROUND Complex PCR applications for large genome-scale projects require fast, reliable and often highly sophisticated primer design software applications. Presently, such applications use pipelining methods to utilise many third party applications and this involves file parsing, interfacing and data conversion, which is slow and prone to error. A fully integrated suite of software tools for...

background: infestation of the skin by the “itch mite” sarcoptes scabiei var. hominis results in a contagious skin infection in humans called “sca-bies”. by resolving morphology issues, the present study was designed to be acquainted with itch mite by molecular markers. methods: the mite samples were collected from scabies patients by visiting government hospitals of twin city, pakistan. for su...

Many protocols in methylation studies utilize one primer set to generate a PCR product from bisulfite modified template regardless of its methylation status (methylation independent amplification MIP). However, proportional amplification of methylated and unmethylated alleles is hard to achieve due to PCR bias favoring amplification of unmethylated relatively GC poor sequence. Two primer design...