Rapid high-yield mRNA extraction for reverse-transcription PCR

Magnetic quantitative reverse transcription PCR: a high-throughput method for mRNA extraction and quantitative reverse transcription PCR.

Over the past few years high-throughput platforms for real-time quantitative PCR have become widely available. The cost of RNA extraction from a large number of samples are, however, quite notable. One method that stands out with respect to free up- or downscaling of sample size and reliability is the isolation of mRNA using oligodeoxythymidylate [oligo(dT)25]-coated magnetic particles. In comb...

One-Step Reverse Transcription PCR (RT-PCR). Fatemeh Mehrpooyan*

Technology for automated, rapid, and quantitative PCR or reverse transcription-PCR clinical testing.

BACKGROUND PCR-based assays can improve clinical care, but they remain technically demanding and labor-intensive. We describe a new instrument, the GeneXpert, that performs automated nucleic acid isolation, reverse transcription, and fluorescence-based quantitative PCR in approximately 35 min. METHODS Yield and integrity of RNA isolated on the GeneXpert were compared with Qiagen-based extract...

mRNA extraction and reverse transcription-PCR protocol for detection of nifH gene expression by Azotobacter vinelandii in soil.

The study of free-living nitrogen-fixing organisms in bulk soil is hampered by the great diversity of soil microbial communities and the difficulty of relating nitrogen fixation activities to individual members of the diazotroph populations. We developed a molecular method that allows analysis of nifH mRNA expression in soil in parallel with determinations of nitrogen-fixing activity and bacter...

Quantification of mRNA using real-time reverse transcription PCR (RT-PCR): trends and problems.

The fluorescence-based real-time reverse transcription PCR (RT-PCR) is widely used for the quantification of steady-state mRNA levels and is a critical tool for basic research, molecular medicine and biotechnology. Assays are easy to perform, capable of high throughput, and can combine high sensitivity with reliable specificity. The technology is evolving rapidly with the introduction of new en...