GSM9630658: Subject 32 PBMC; Homo sapiens; RNA-Seq
Aspirated alveolar fluid and blood were collected in parallel from patients in cell preparation tubes (CPT) and immediately processed. Cells were cryopreserved and later thawed for RNA isolation. Total RNA was extracted from BAL cells and PBMCs using standard column-based RNA extraction methods according to manufacturer instructions. BAL (AmpliSeq): Targeted RNA libraries were prepared using the Ion AmpliSeq Human Transcriptome Panel. Sixteen nanograms of total RNA was reverse transcribed, amplified, barcoded, purified with magnetic beads, and PCR-amplified per manufacturer protocol. Libraries were quantified, pooled, templated on an Ion Chef Instrument, and sequenced on an Ion Torrent S5 Prime.  PBMC (RNA-seq): Libraries were prepared using the KAPA mRNA HyperPrep kit. Poly(A)+ mRNA was isolated from 50 ng total RNA, fragmented, converted to cDNA, adapter-ligated with dual indices, PCR amplified, pooled, and sequenced on an Illumina NovaSeq 6000 (paired-end).