GSM5328010: 278_Tvm_pan-KIR_positive; Homo sapiens; RNA-Seq
FACS- sorted TNAIVE, TEMRA, KIR+RA+, and NKG2A+RA+ T- cell sub-sets of six healthy donors (ages: 30, 57, 61, 63, 65, 66 years) were centrifuged at 485 g for 15 min and lysed in buffer RLT (Qiagen)   containing 1% β- mercaptoethanol. Total RNA was extracted from the cell lysates by using the RNeasy kit (Qiagen) according to the manufacturer's protocol. RNA integrity was assessed by using the RNA 6000 Pico kit (Agilent Technologies) on a 2100 BioAnalyzer (Agilent Technologies). All RNA Integrity Number (RIN) scores were >7 DNA synthesis and amplification were performed by using the SMART- Seq® v4 Ultra® Low Input RNA kit for sequencing (Takara) and AMPure XP beads (Beckman Coulter) were used to purify the samples. Proper size distribution of the fragments acquired was veri-fied using the High Sensitivity DNA kit (Agilent Technologies). Next, 1 ng of cDNA was used to prepare a Nextera XT DNA library accord-ing to the manufacturer's instruction (Illumina). Libraries were sub-sequently validated for fragment size using QIAxcel DNA Screening Kit (Qiagen) and quantified using RT- qPCR with a KAPA Library Quantification kit (KK4824, Roche/KAPA Biosystems). 23 librar-ies were pooled at equimolar concentrations and sequenced using the Illumina NextSeq 500/550 High Output Kit v2.5 (single- end, 75 cycles)