Bio-Rad Laboratories has released the Iscript RT-qPCR Sample Preparation Reagent for rapid isolation of total RNA and enabling reverse transcription and real-time PCR directly from cell lysates.
Scientists can use the protocol to efficiently remove genomic DNA and stabilise RNA in as little as five minutes.
Viresh Patel, senior product manager, PCR reagents, Bio-Rad Laboratories, said: '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.' 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 stabilisation, and genomic DNA removal for sensitive, quantitative gene-expression analysis.
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.
Patel added: 'The cost of RNA sample preparation has remained quite high, relative to the cost of performing reverse transcription and quantitative PCR.
'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 stabilises 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 -20C.
The reagent is compatible with multiple PCR reagents already being used by researchers.
