GSM9041578: Human CALB2-expressing neuron 8498_S8 transfected with CellREADR; Homo sapiens; RNA-Seq
Clean nucleus extraction was confirmed visually with both DIC and fluorescent images. Nuclei were stored at -80C in 10μL of lysis buffer with RNase inhibitor (40 U/µL) and then processed for sequencing in batches (5-12 nuclei/batch). Nuclear RNA was processed into cDNA libraries using the SMART-Seq v4 Ultra Low Input kit (Takara) using 19 PCR cycles. Nuclei that yielded >200 pg/µL cDNA were sequenced. Borderline cDNA (concentrations just below 200 pg/µL, and fragment sizes >500 bp) were re-amplified with 3 rounds of PCR, rechecked for cDNA quality, and included in the sequencing set if reamplification increased cDNA concentration above threshold.