ReliaPrep™ RNA Miniprep Systems

Protocol tips

Upstream tips	Protocol tips	Downstream tips
- Before beginning the ReliaPrep™ RNA Cell Miniprep System protocol, four solutions must be prepared.

Upstream tips
- Before beginning the ReliaPrep™ RNA Cell Miniprep System protocol, four solutions must be prepared.

Publication protocol

Total RNA was then isolated from the cell cultures with the ReliaPrep RNA Cell Miniprep System following the manufacturer's instructions. Pre-designed and validated gene-specific primers (QuantiTect Primer assay) were used with the GoTaq One-Step qRT-PCR System assay. PCR efficiency was verified to be between 90-105% under our conditions and products generated gave a single melt peak in post-run analysis. The following PCR protocol was used: a 15 min cycle at 37°C for reverse transcription followed by a 10 min denaturation at 95°C and 35 cycles of 10 sec denaturation at 95°C, followed by a 30 sec annealing at 60°C and an extension of 30 sec at 72°C. The PCR reaction volume was 20 μl. GAPDH was used as a housekeeping gene to normalize data and wells without reverse transcriptase were used as a control for genomic DNA. Fold change data was calculated by the 2−ΔΔCt method. Samples were run in triplicate and three biological replicates were obtained. Samples were analyzed using a Mastercycler ep Realplex (Eppendorf, Hauppauge, NY, USA) using the Realplex 1.5 software (Eppendorf, Hauppauge, NY, USA).

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Manufacturer protocol

Download the product protocol from Promega for ReliaPrep™ RNA Miniprep Systems below.

Download manufacturer protocol

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