Mrna cdna synthesis kit

Total RNA was extracted from cell lines using the AG RNAex Pro reagent (Accurate Biology). 2 μg total RNA was used to synthesized single-stranded cDNA by All-in-OneTM miRNA or mRNA cDNA synthesis kit (GeneCopoeia Inc., MD, USA). qRT-PCR was carried out on the ABI QuantStudio 7 Flex Real-Time PCR System using All-in-One Kit (GeneCopoeia Inc., MD, USA) and sets of gene-specific primers. The reaction parameters included an initial step at 95 °C for 10 min, 40 cycles of 95 °C for 10 seconds, 62.5 °C for 20 seconds, and 72 °C for 15 seconds. U6 and GAPDH were used as endogenous control for miRNA and mRNA, respectively, and data were gauged through the classical 2^-ΔΔCt method. Sequences of primers used in this study were listed in Table S1.

Total RNA was isolated from cell lines using the Trizol reagent (Invitrogen, CA, USA), and first strand cDNA was generated using the All-in-One^TM miRNA or mRNA cDNA synthesis kit (GeneCopoeia Inc., MD, USA) in a 25 μL reaction system containing 1 μg of total RNA. A 0.5 μL aliquot of cDNA was amplified using All-in-One^TM miRNA or mRNA Mix (GeneCopoeia Inc., MD, USA) in each 20 μL reaction system. The amplification procedure was performed using the ABI 7300 Fast Real-Time PCR system (Applied Biosystems), and the reaction parameters included an initial step at 95°C for 10 min, 40 cycles of 95°C for 10 seconds, 62.5°C for 20 seconds, and 72°C for 15 seconds. Expression values were calculated using the 2^-ΔΔCT method and normalized to an internal control (U6 for miRNA and GAPDH for mRNA). The primers used for the qRT-PCR are listed in Supplementary Table S1.