Bulge loop hsa mir 93 5p qrt pcr primer set

Total RNA from lung tissues or in vitro cultured hBSMCs was extracted with TRIzol reagents (Invitrogen, CA, USA) according to the manufacturer’s instructions. For Orai1 and H19, RNA was reversely transcribed into cDNA using a ReverTra Ace qPCR RT kit (Toyobo, Tokyo, Japan). cDNA was then amplified with SYBR Premix Ex Taq™ II (RR820A, Takara, Japan) via the following three steps: 1 cycle at 95°C for 30s, followed by 40 cycles of 95°C for 5s and 60°C for 30s, and a melting curve collected at 95°C for 5 s and 60°C for 1 min. For miR-93-5p detection, reverse transcription and qRT-PCR were performed using the Bulge-Loop^TM miRNA qRT-PCR starter kit (R11067.2, RiboBio Co., Guangzhou, China) and the Bulge-Loop^TM hsa-miR-93-5p qRT-PCR primer set (R10031.7, RiboBio Co., Guangzhou, China). The amplification process included 1 cycle at 95°C for 10 min, 40 cycles of 95°C for 2 s, 60°C for 20s, and 70°C for 10s. The cycle threshold (CT) values of miR-93-5p were normalized to U6, and CT values of H19 or Orai1 were normalized to β-actin for respectively. The reaction was performed using the CFX96 Real-Time system (Bio-Rad, USA). Data were analyzed using 2^−ΔΔCT method.