Total RNA was extracted from cells using an RNeasy Mini Kit (Qiagen). cDNA was synthesized and real-time quantitative RT-PCR was performed using the SuperScript III Platinum SYBR Green One-Step quantitative RT-PCR Kit (Thermo Fisher Scientific). The quantification of gene expression was performed in triplicate. Amplification of the sequence of interest was normalized with a housekeeping gene, ACTB. Quantification was performed using comparative threshold cycle (Ct) method of analysis. The value was expressed as a fold change relative to RNA from GFP-expressing or untreated control cells. Intron-spanning quantitative PCR primers were designed using the assay design center of the Universal ProbeLibrary (Roche).
Superscript 3 platinum sybr green one step quantitative rt pcr kit
Manufactured by Thermo Fisher Scientific
The SuperScript III Platinum SYBR Green One-Step quantitative RT-PCR Kit is a reagent kit for performing one-step reverse transcription and real-time PCR amplification using SYBR Green detection. It is designed for the quantification of RNA targets.
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2 protocols using superscript 3 platinum sybr green one step quantitative rt pcr kit
1
Quantitative RT-PCR Analysis of Gene Expression
2
Quantification of Gene Expression
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Total RNA was extracted from cells using RNeasy Mini Kit (Qiagen). cDNA was synthesised and real‐time quantitative RT–PCR was performed using the SuperScript III Platinum SYBR Green One‐Step quantitative RT–PCR Kit (Thermo Fisher Scientific). The quantification of gene expression was performed in triplicate. Amplification of the sequence of interest was normalised with a housekeeping gene, PUM1. Quantification was performed using comparative threshold cycle (Ct) method of analysis. The value was expressed as a fold change relative to RNA from proliferating cells.
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