GSM9981158: Infected with 1T, 24hr, rep 3; Homo sapiens; RNA-Seq
Macrophages were lysed using a previously described RLT buffer-based protocol. Following DNA removal using RNase-free DNase, RNA samples were subjected to purification using RNeasy kit (QIAGEN) per manufacturer's instruction. High quality RNA samples determined by running RNA samples on 1.5% agarose gel and NanoDrop (ThermoFisher) The RNA sequencing libraries were prepared using the NEBNext Ultra II RNA Library Prep Kit for Illumina according to the manufacturer's instructions (New England Biolabs, Ipswich, MA, USA). Briefly, mRNAs were initially enriched with oligo(T) beads. Enriched mRNAs were fragmented for 15 min at 94°C. First- and second-strand cDNA was subsequently synthesized. cDNA fragments were end repaired and adenylated at the 3′ ends, and universal adapters were ligated to cDNA fragments, followed by index addition and library enrichment by PCR with limited cycles.