Total RNA was isolated using TRIzol® Reagent (Life Technologies, 15596-018) according to manufacturers’ instructions. Subsequently, complementary DNA was generated using the PrimeScript™ RT reagent Kit with gDNA eraser (Takara Bio Inc., RR047A), and quantitative real-time reverse transcription polymerase chain reaction (qPCR) was performed using the Real-Time PCR detection system (Applied Biosystems, 7500) with 2× SYBR Green II/ROX qPCR Master Mix (Takara Bio Inc., RR82LR). Relative mRNA expression was calculated using the delta threshold cycle (ΔΔCT) method and normalized to β-actin (ACTB) expression. The PCR primers are listed in Additional file: Table S2 .
Sybr green 2 rox qpcr master mix
Manufactured by Takara Bio
Sourced in China
The 2× SYBR Green II/ROX qPCR Master Mix is a ready-to-use solution for quantitative real-time PCR (qPCR) reactions. It contains SYBR Green II dye, which binds to double-stranded DNA, and ROX passive reference dye for signal normalization.
Automatically generated - may contain errors
2 protocols using sybr green 2 rox qpcr master mix
1
Quantitative RT-qPCR Analysis of Gene Expression
2
RNA Isolation and qPCR Analysis
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Total RNA was isolated using TRIzol® Reagent (#15596-018, Life Technologies, New York, USA) according to manufacturers’ instructions. Subsequently, cDNA was generated using the PrimeScript™ RT reagent Kit with gDNA eraser (Takara Bio Inc., Dalian, China), and quantitative real-time reverse transcription polymerase chain reaction (qPCR) was performed using the Real-Time PCR detection system (#7500, Applied Biosystems, Shanghai, China) with 2× SYBR Green II/ROX qPCR Master Mix (Takara Bio Inc.). Relative mRNA expression was calculated using the delta threshold cycle (ΔΔCT) method and normalized to β-actin (ACTB) expression. The PCR primers are listed in Additional file 1 : Table S1.
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