Hairpin it micrornas quantitation kits

Relative expression levels of lncRNAs and miRNAs in RA FLS were determined by real-time PCR (26 (link)). Total RNA was isolated by a TRIzol reagent and quantified with NanoDrop ND-2000 (Thermo Scientific). Quality control was performed by Agilent Bioanalyzer 2100 (Agilent Technologies Inc.). The RNA was reverse transcribed by a PrimeScript^TM RT reagent kit containing a gDNA eraser (Takara). Real-time PCR was conducted with a SYBR Premix Ex TaqTM kit (TliRNaseH Plus) in an Mx3005P qPCR System (Agilent Technologies Inc.). Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was used as an internal control. The primers used were listed below: WAKMAR2: 5′-GGCCTCAGTGAGGTAAATCG-3′; 5′-CATACCACTACACTCCAGC-3′ and GAPDH: 5′-AACTTTGGCATTGTGGAAGG-3′; 5′-GGATGCAGGGATGATGTTCT-3′. The expressions of miRNAs were detected by stem-loop real-time PCR. The miRNAs from total RNA were reverse transcribed and subjected to amplification using Hairpin-it™ MicroRNAs Quantitation kits (GenePharma), which contained both primers of miRNAs and the internal control U6. The obtained data were analyzed by the MXProv 4.1 Sequence Detection System and calculated according to the 2^−ΔΔCt formula (27 (link)).