CellsDirect™ Two-Step qRT-PCR Kit - FAQs

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2 product FAQs found

What is the difference between one-step and two-step RT-PCR?

One-step RT-PCR allows easier processing of large numbers of samples and helps minimize carry-over contamination since tubes are not opened between cDNA synthesis and amplification. By amplifying the entire cDNA sample, one-step RT-PCR can provide greater sensitivity-down to 0.01 pg total RNA. Only gene-specific primers can be used with one-step kits.

Two-step RT-PCR is useful for detecting multiple messages from a single RNA sample. You have greater flexibility to choose polymerase and primers than you do with one-step RT-PCR systems. When performing two-step RT-PCR you have the option of using either oligo(dT), random hexamers, or gene-specific primers, and then performing PCR in combination with either Platinum Taq DNA Polymerase, Platinum Taq DNA Polymerase High Fidelity, or your choice of other PCR enzymes.

I want to perform qRT-PCR using cDNA that I synthesize directly from cells. Can I use the SuperScript III CellsDirect cDNA Kit for end-point PCR and the Platinum Quantitative PCR SuperMix-UDG kit?

We would recommend using our Cells-to-CT kits for best qRT-PCR results. The CellsDirect kits for end-point PCR have not been optimized for qRT-PCR.