title: GSM7865566: BFCN, Ts65Dn, Treatment, 15-36; Mus musculus; RNA-Seq
library_construction_protocol: 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 antibody detection method for ChAT positive neurons.  ChAT-IR neurons were isolated by laser capture microdissection (LMD7000) and stored in Qiazol at -80*C until ~500 neurons per sample were collected.  RNase-free precautions were employed throughout, and solutions were made with 18.2 mega Ohm RNase-free water.   RNA from ~500 BFCNs was purified using miRNeasy Micro kit (Qiagen) following the manufacturer's specifications, with a the optional DNase digestion step done twice sequentially. RNA libraries were prepared by using SMARTer Stranded Total RNA-Seq kit-Pico input Mammalian with minor modifications to utilize full volume of RNA. RNA-seqSamples were quantified (Agilent 2100 HS DNA kit), and samples below 2 nM of library were excluded. Samples were pooled in equimolar concentrations and assayed on an Illumina HiSeq-4000 using a single read 50 cycle protocol at the NYUGSOM Genome Technology Center.