Abi 7300 thermal cycler

Total RNA was extracted using TRIzol reagent (Invitrogen), and complementary DNA was synthesized using a PrimeScript RT kit (TaKaRa, Shiga, Japan). The real-time PCR analysis was performed using an ABI 7300 thermal cycler (Applied Biosystems, Foster City, CA, USA) with SYBR Green Q-PCR Master Mix (TaKaRa). The ΔΔCt method was used to calculate the relative amount of mRNAs with the 36b4 as an internal control. The primer sequences are listed in Table 1.

Total RNA was extracted from the cells with TRIzol (Invitrogen Life Technologies) and reverse transcribed into cDNA with reverse transcriptase from the Primescript RT Reagent kit (Takara Biotechnology Co., Ltd., Dalian, China), as described previously (21 (link)). The RT-qPCR reactions were performed using the Thermal Cycler Dice Real-Time PCR system (Takara Biotechnology Co., Ltd.) and the ABI 7300 Thermal Cycler (Applied Biosystems Life Technologies, Carlsbad, CA, USA) with the following cycling conditions: Initial denaturation at 95°C for 10 min, and 40 cycles of 95°C for 30 sec, 55°C for 30 sec, and 72°C for 2 min. In order to ensure the reproducibility of the results, all the genes were tested in triplicate. The following primers from BGI Tech (Shenzhen, China) were used for the present study: ITGB1, sense 5′-GTCTCAGACTGGCTTCAGTG-3′, antisense 5′-ATGATTGTACCGAGGCTGTC-3′; FAK, sense 5′-GCGGCCCAGGTTTACTGAA-3′, antisense 5′-GGCCTGTCTTCTGGACTCCAT-3′; and β-actin (human), sense 5′-AGGGGCCGGACTCGTCATACT-3′, and antisense 5′-GGCGGCACCACCATGTACCCT-3′. Reaction of each sample was performed in triplicate. Relative expression levels were calculated using the 2-∆∆Ct method.