Accupower cdna synthesis kit

Total RNA was extracted from MSCs using the EasyBlue RNA isolation reagent (Intron Biotechnology, Sungnam, Korea). cDNAs were synthesized from 2 μg total RNA using the AccuPower cDNA synthesis kit (Bioneer, Daejeon, Korea). Semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) was performed using the AccuPower PCR premix (Bioneer) in a thermal cycler (Bio-Rad C1000; Bio-Rad, Hercules, CA, USA). The amplified PCR products were electrophoresed on 1% agarose gels containing SybrSafe (Invitrogen, Carlsbad, CA, USA) and analyzed using a fluorescence image analyzer (LAS4000 mini; Fujifilm, Tokyo, Japan). PCR was performed with the following primers: IL-10 (forward 5′-ATCCAAGACAACACTACTAA-3′ and reverse 5′-TAA ATATCCTCAAAGTTCC-3′; product 587 bp) and glyceraldehyde 3-phosphate dehydrogenase (GAPDH; forward 5′-CCACTGGCGTCTTCACCAC-3′ and reverse 5′-CCTGCTTCACCACCTTCTTG-3′; 476 bp). To confirm the expression of ICOSL mRNA, quantitative RT-PCR (qPCR) was performed using the ICOSL TaqMan assay (Assay ID: Hs00323621_m1; Applied Biosystems, Waltham, MA, USA) and the TaqMan Universal PCR Master Mix (Applied Biosystems). The mRNA expression level was normalized to 18S rRNA gene expression (Hs03928985_g1) as an internal control. The qPCR was run on the StepOnePlus real-time PCR System (Applied Biosystems).

The mRNA expression of cytokines in nasal tissue and HNECs was examined using qRT-PCR, as previously described [15 (link)]. Briefly, total RNA was extracted using an easy-BLUE reagent (Intron Biotechnology, Seongnam, Republic of Korea), followed by cDNA synthesis from 2 μg of total RNA using the AccuPower cDNA synthesis kit (Bioneer, Daejeon, Republic of Korea). The cDNA was amplified and quantified on a PCR system using SYBR Green Master Mix (Applied Biosystems, Foster City, CA, USA). The primer sequences used to amplify the specific genes are listed in Supplementary Table S1.