Endpoint genotyping software

Genotyping of IFNL3 single-nucleotide polymorphism rs12979860 and the IFNL4-ΔG SNP, rs368234815, were conducted using custom TaqMan^® SNP Genotyping Assays from Applied Biosystems (Life Technologies, Carlsbad, CA, USA) performed according to manufacturer’s instructions. The Roche LightCycler 480 Real-Time System (Roche Applied Science, Indianapolis, IN, USA) was used to amplify the DNA and measure the fluorescence from each reaction to determine each genotype as calculated by Endpoint Genotyping Software (Roche, Nutley, NJ, USA).
Human IFNL protein levels have been challenging to measure and discriminate between the different subtypes: human IFNL1/IFNL3 levels were measured using a commercially available EIA kit (R&D Systems Inc., Minneapolis, MN, USA; Cat. number DY1598B).

The rs8099917 and rs12979860 polymorphisms were determined using the Invader-Plus assay [18 (link)], which combines PCR and the Invader reaction [19 (link), 20 (link)], on a LightCycler 480 (Roche Diagnostics, Basel, Switzerland). The Invader-Plus assay reagent kit, purchased from Third Wave Technology (Madison, USA), consists of a probe mix, a buffer mix, and an enzyme mix. The reagents were premixed according to the manufacturer’s instructions. Then, 10 ng of genomic DNA was added to the master mix. The primer and probe sets are described in Table S1. The cycle conditions were 18 cycles of 15 s at 95 °C and 60 s at 70 °C. At the end of the PCR, the Taq polymerase was inactivated at 99 °C for 10 min and the reaction temperature was lowered to 63 °C for 15–30 min to permit the hybridization of the probe oligonucleotide and the formation of the overlap flap structure. Data were analyzed by endpoint genotyping software (Roche Diagnostics). Both rs8099917 and rs12979860 were determined from the African-American samples; however, we tested the Japanese samples for only rs8099917 because it was previously reported that rs8099917 and rs12979860 represented 98.6 % of the Japanese population [18 (link)].