GSM4473697: Plate_Q5; Mus musculus; RNA-Seq
For each sample, individual labelled cells were isolated by manual cell picking on a fluorescence dissecting scope, after incubating slices in ACSF with neuronal activity blockers (TTX, APV, DNQX) and pronase for 50 minutes. Each isolated cell was transferred to a 0.2mL PCR 8-tube strip containing 3 microliters of lysis buffer (0.2% Triton X-100) in Nuclease-free water with RNase inhibitor. Each 8-tube strip was flash frozen on dry ice and kept at -80C until library prep and sequencing. Each single cell underwent the RT process, and cDNA was then pooled before amplification, using the SMART-SCRB technique (using SMARTer-based amplification with cell barcodess). Amplified cDNA was fragmented to ~250bp, ligated to Illumina sequencing adaptors with Encore Rapid kit, and sequenced on an Illumina HiSeq 2500 with 4-fold multiplexing (paired end, 25bp for Read 1 and 125bp for Read 2)