QuantiTect SYBR Green PCR Kit (1000)

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Publication protocol

RNA Isolation and qRT-PCR: For detection of PANTR1 and other gene expression levels in ccRCC cells (confluency of approximately 75–90%) and human tissue, total RNA was isolated using a standard Trizol (Invitrogen, Carlsbad, CA, USA) protocol according to the manufacturer’s instructions. RNA was stored at −80 °C until further processing. cDNA was synthesized from 1 µg of total RNA using the QuantiTect Reverse Transcription Kit (Qiagen, Venlo, The Netherlands). Quantification of lncRNAs and mRNAs was performed using the QuantiTect SYBR Green PCR kit (Qiagen) according to the manufacturer’s recommendations on a LightCycler 480 real-time PCR system (Roche, Mannheim, Germany). U6 (Eurofins Scientific, Vienna, Austria) was used for normalization of lncRNAs and GAPDH (Eurofins) for protein coding genes. Differences in fold expression were calculated in Excel (Microsoft Corporation, Redmond, WA, USA) following the 2−ΔΔCt method. All specific primer sequences for analyzed genes are listed in Table S1. Detailed experimental performance according to MIQE guidelines are provided in Supplementary Text 1.

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Manufacturer protocol

Download the product protocol from Qiagen for QuantiTect SYBR Green PCR Kit (1000) below.

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