Type it fast snp probe pcr master mix

Informed consent for genetic analysis was obtained from all patients. Genomic DNA was extracted from buffy coat using Maxwell RSC Buffy Coat DNA Kit (Promega) as per the manufacturer's protocol and quantified using Qubit (Thermofisher). DNA samples from patients were genotyped using the Affymetrix UK Biobank array, as described elsewhere (Ansari et al., 2017 (link)). Phasing and imputation was performed using SHAPEIT2 (Delaneau et al., 2013 (link))Dao Thi et al., 2011 (link) and IMPUTE2 (Howie et al., 2009 (link)) version 2.3.1 using default settings and the 1000 Genomes Phase III dataset as a reference population (Auton et al., 2015 (link)). Imputation quality was high for both rs117648444 and rs368234815 variants (information 0.974 and 0.994 respectively and certainty 0.995 and 0.997 respectively). Patients from the EAP cohort from whom enough DNA was available (62/74) were also independently genotyped for both rs117648444 and rs368234815 variants. Genotyping of IFNL4 rs368234815 and rs117648444 was performed on DNA using the TaqMan SNP genotyping assay and sequences described previously (Prokunina-Olsson et al., 2013 (link)) with Type‐it Fast SNP Probe PCR Master Mix (Qiagen). The concordance between genotyped and imputed genotypes was 100% for both variants.

Total RNA was reverse‐transcribed with the SuperScript^® VILO^™ Kit (Life Technologies, Paisley, UK), and gene expression was measured by quantitative real‐time PCR (Q‐PCR) using TaqMan^® assays (Life Technologies; Table S1). Statistical significance was calculated using the Kruskal–Wallis test in GraphPad Prism 5 (GraphPad Software, La Jolla, CA, USA). Genotyping of IFNL4 rs368234815 was performed on cDNA using the assay described previously 18 with Type‐it Fast SNP Probe PCR Master Mix (Qiagen).