GSM7220677: 110653_3_S82; Homo sapiens; RNA-Seq
In DAISY, venous blood samples are collected at 9, 15, and 24 months and yearly thereafter. Beginning in November of 2007, RNA samples were collected from all DAISY participants with Tempus tubes (Applied Biosystems). We selected samples from two visits (termed IA-1 and IA-2) that occurred after the onset of IA for mRNA sequencing. The IA-1 visit was selected as the first available visit after the onset of IA for all groups. In the progressor group, the visit that preceded clinical onset of T1D was selected as the IA-2 visit. In the maintainer and reverter groups, the IA-2 visit was selected as the visit where age was closest to median age in the progressor group at the IA-2 visit. Total RNA was isolated and purified from frozen (−80°C), peripheral whole blood samples ofn= 138 subjects meeting the inclusion criteria (Supplementary Material). We also included 12 control samples and 12 technical replicates. NuGEN library preparation kits were used to construct the strand-specific total RNA libraries. The Globin AnyDeplete kit was used to remove globin transcripts. RNA integrity number (RIN) scores for all samples were quantitated with use of the TapeStation (Agilent). Paired-end 150-cycle 2 × 150 sequencing was performed with the Illumina NovaSeq 6000 system. Samples were pooled together in a single flow cell and were sequenced twice to obtain the desired read depth, resulting in a median read depth of 90 million paired end reads across the samples (range 54–128 million).