title: GSM3137040: N719_S511_2: CD69pos scRNA-seq; Homo sapiens; RNA-Seq
library_construction_protocol: scRNA-seq: Single-cell analysis was derived from a previously published method (Shalek et al., Nature, 2014). Single HIV-tetramer+ cells were index sorted using a FACSAria II (BD Biosciences) directly into 96-well microtiter plates containing lysis buffer. In total, 552 cells were sorted. Cellular nucleic acids were recovered by processing the lysates with an RNEasy Plus Micro Kit (Qiagen) and then binding to RNAClean (SPRI) beads (Beckman Coulter). ERCC RNA control (Ambion) was diluted 1 in 107 from stock, and 10% by volume was added to the RNAClean beads prior to purification. scRNA-seq: Libraries were prepared by getting mRNA be reverse transcribed from an anchored oligo-dT primer that includes a universal priming site at the 5' end. RT reactions also included a template-switch oligonucleotide to allow the incorporation of the universal priming site at cDNA 3' ends. Whole transcriptome cDNA was then amplified by PCR for 22–24 cycles using universal priming sites primers and Kapa HiFi Hot Start ReadyMix. After post-PCR clean-up, amplified whole transcriptome cDNA was incorporated into barcoded Illumina Nextera libraries and sequenced to a depth of about 2 million 150bp paired-end reads/cell on a HiSeq 4000 (Illumina).  and amplified for 14 PCR cycles. Libraries were pooled and paired-end sequenced (38bp+37bp).
attributes.tag: GEO Accession
attributes.value: GSM3137040