GSM8669896: 15-36-DG, Ts65Dn, ChP; Mus musculus; RNA-Seq
Brain tissues were immediately flash frozen and 20 μm-thick tissue sections were cryostat cut (−25°C) in the coronal plane and mounted on polyethylene naphthalate (PEN) membrane slides. Sections were stained with rapid Nissl staining protocol. CA1 and CA3 PNs along with DG GCs were identified visually in the same section and isolated by laser capture microdissection (LCM). Approximately 6 rostral hippocampi sections were dissected by LCM (approximately -1.58 to -2.54 Bregma), with mean collections of ~950 CA1, ~750 CA3 and ~1175 DG neurons collected per brain before proceeding to RNA isolation and RNA-seq library preparation. RNA from each sample was purified using miRNeasy Micro kit (Qiagen) following the manufacturer's specifications, with a the optional DNase digestion step done twice sequentially. The SMARTer Stranded Total RNA-Seq kit-Pico input Mammalian v3  was employed with unique indexes for each sample. To normalize input, CA1, CA3 and DG RNA concentrations were used to determine sample input, with average of ~3-3.75 ng RNA input utilized per sample. Samples were fragmented for 3.75 min, with 5 cycles for PCR1 and 14 cycles for PCR2. Samples were quantified using Tapestation D1000 DNA assay.  Samples were pooled in equimolar concentrations and assayed on an Illumina NovaSeq 6000 using a S1 100cycle flow cell (v1.5) by the NYUGSOM Genome Technology Center.