Truseq dual index sequencing primers

High quality DNA was selected for NGS sequencing. We performed targeted panel sequencing to identify genetic variants within the UCP1 gene for MetS patients and controls. To build amplicon sequencing libraries for the coding region of the UCP1 gene, we used TruSeq Custom Amplicon Low Input Kit (Illumina, San Diego, CA, USA) with TruSeq Dual Index Sequencing Primers (Illumina, San Diego, CA, USA). For 3′UTR and 5′UTR regions of the UCP1 gene, we used AmpliSeq Illumina Custom DNA Panel (Illumina, San Diego, CA, USA) with AmpliSeq CD Indexes (Illumina, San Diego, CA, USA). The quality and quantity of the purified libraries was assessed by Qubit 3.0 Fluorometer (ThermoFisher Scientific, Waltham, MA, USA) using Qubit dsDNA HS Assay (ThermoFisher Scientific, Waltham, MA, USA) and TapeStation 4200 (Agilent Technologies, Santa Clara, CA, USA) using High Sensitivity D1000 ScreenTape Assay (Agilent Technologies, Santa Clara, CA, USA). Only high-quality libraries were sequenced on the MiSeq System (Illumina, San Diego, CA, USA) using the V2 Illumina Sequencing Kit 500 (Illumina, San Diego, CA, USA).

Single-cell RNA sequencing was performed as described by Picelli
et al.^{10 (link)}. Sequencing libraries were prepared using the TruSeq dual-index sequencing primers (catalog no PE-121-1003, Illumina, San Diego, California) and paired-end sequencing was performed on the Illumina HiSeq4000 platform. All samples were spiked with ERCC RNA Spike-In Mix (catalog no 4456740, Thermo Fisher) as an internal control for sequencing (Extended data
^{11 (link)}, Figure S1D).