title: GSM8161058: Multiplexed sublibraries 2 and 6 [ob/ob male; Sample 13,14,19, and 22]; Mus musculus; RNA-Seq
library_construction_protocol: Wild type (C57BL/6J) and ob/ob male mice were fed a standard purified diet AIN-93M, for 8 wks. Mice were sacrificed at 18 weeks of age and hippocampus was dissected from left hemisphere and then stored at -80 degree celcius. Hippocampal nuclei was isolated and fixed using Parse biosciences nuclei isolation and fixation protocol. Barcoded single cell libraries were prepared from fixed single cell suspensions using Evercode Whole Transcriptome Mega kit (Parse Biosciences) following the manufacturer's instructions.  Individual transcriptomes from all samples were uniquely labeled by passing fixed nuclei through four rounds of barcoding. In each round, pooled cells are randomly distributed into different wells, and transcripts are labeled with well-specific barcodes. Using next-generation sequencing, cells from individual samples are identified using the first well (first barcode) placement. Each sublibrary contains uniquely barcoded cells from all samples.  At the end of library prep, cells are distributed into 16 different sublibraries. The cDNA and library fragment size distribution were verified on a Bioanalyzer 2100 and TapeStation, respectively. The libraries were quantified by fluorometry on a Qubit instrument and by qPCR with a Kapa Library Quant kit prior to sequencing. The libraries were sequenced with paired-end 100 bp reads.  The sequencing generated approximately 35,000 reads per cell.