title: GSM4799170: NK_post ketogenic diet [MUC11709_S28]; Homo sapiens; RNA-Seq
library_construction_protocol: After stimulation, CD3/CD28 Dynabeads were magnetically removed. Pan T cell-, CD4+- and CD8+-cell-isolation was performed by magnetic cell separation (Pan T Cell Isolation Kit, # 130-096-535 | human CD4 MicroBeads, # 130-045-101 | human CD8 MicroBeads, # 130-045-201, Miltenyi Biotec, Bergisch Gladbach, Germany) using an AutoMACS Pro Separator ( # 130-092-545, Miltenyi Biotec, Bergisch-Gladbach, Germany) according to the manufacturer's instructions. Total RNA was isolated from primary T cells using the mirVana miRNA Isolation Kit (Thermo Fisher Scientific, Waltham, MA, USA) with subsequent DNase treatment (Turbo DNase, Thermo Fisher Scientific, Waltham, MA, USA) or miRNeasy RNA Isolation Kit (#217004, Quiagen, Hilden, Germany) with on-column DNA digestion. For RNA purification, the Monarch RNA Clean-up Kit (New England Biolabs, Ipswich, MA, USA) was utilized. All isolation procedures have been performed according to the respective manufacturer's instructions. RNA quantity and purity were measured using a NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific, Waltham, MA, USA). First-strand cDNA was synthesized from equal amounts of RNA using Superscript III reverse transcriptase (Invitrogen, Carlsbad, CA, USA), random hexamers and oligo(dT) primers. RNA libraries were prepared for sequencing using standard Illumina protocols
attributes.tag: GEO Accession
attributes.value: GSM4799170