Bio-Rad’s iScript RT-qPCR Sample Preparation Reagent Provides Reverse Transcription-Ready RNA in Five Minutes
Product News Nov 04, 2009
Bio-Rad Laboratories, Inc. has announced the availability of its iScript™ RT-qPCR Sample Preparation Reagent for rapid isolation of total RNA and enabling reverse transcription and real-time PCR to be performed directly from cell lysates.
Scientists can use the protocol to efficiently remove genomic DNA and stabilize RNA in as little as five minutes.
“We designed iScript RT-qPCR Sample Preparation Reagent to meet the needs of researchers performing high-throughput gene expression analysis, gene silencing (or RNAi), and microarray validation,” said Viresh Patel, Senior Product Manager, PCR Reagents, Bio-Rad Laboratories.
Conventional RNA purification methods, such as organic extraction or column-based methods that include salting out or the use of magnetic particles, require numerous steps to purify RNA without contaminating genomic DNA and can take at least 30 minutes to complete. iScript RT-qPCR Sample Preparation Reagent employs selective lysis of the cell membrane while leaving the nuclear membrane intact to achieve DNA-free RNA within five to 10 minutes.
The single buffer formulation contains all the components needed to perform cell lysis, RNA stabilization, and genomic DNA removal for sensitive, quantitative gene expression analysis, and it offers sensitive detection of high-, medium-, and low-copy gene targets directly from cell lysates. It also enables multiplex real-time detection of up to four targets from as few as 10 cells.
“The cost of RNA sample preparation has remained quite high, relative to the cost of performing reverse transcription and quantitative PCR,” said Patel. “This reagent provides a more affordable option for researchers looking to complete sensitive, high-throughput gene expression analysis.”
Upon exposure of cells to iScript RT-qPCR Sample Prep Reagent, cell disruption and lysis occurs in approximately 30 seconds with the aid of mild vortexing. The reagent also stabilizes RNA and effectively inhibits intracellular ribonucleases (RNases). A short two-minute high-speed centrifugation yields total RNA that can be collected in the supernatant while the nuclei containing genomic DNA form a pellet and can be discarded. The resulting RNA preparation is ready for use, without further treatment, in subsequent reverse transcription and PCR/qPCR.
RNA isolated with the iScript RT-qPCR Sample Preparation Reagent is stable for up to six months when stored at -20°C. The reagent is compatible with multiple PCR reagents already being used by researchers.