GSM7719456: 5xFAD;Bmp4-cKO mice, cortex, replicate 4; Mus musculus; RNA-Seq
Mice were deeply euthanized by CO2 asphyxiation and subsequently transcardially perfused with ice-cold PBS, pH 7.4. The brain tissues were dissected, and the cortical tissues were collected. The tissues were cut into 1mm slices and then dissociated enzymatically by 1 mg/ml collagenase type II (Worthington, 4176) and 2 mg/ml DNase 1 (Sigma-Aldrich, DN25) in 20 ml neurobasal media (Gibco, 21103049) at 37°C for 30 min.  Tissue slices were pelleted by centrifugation at 300g for 10 min and dissociated once more in a papain solution [1 mg/ml papain (Worthington, LS003119) and 2 mg/ml DNase 1 (Sigma-Aldrich, DN25) in 20 ml neurobasal media (Gibco, 21103049)] at 37°C for 30 min. The enzymes were then inactivated by adding 20 μg/ml ovoicoid protease inhibitor (OIA) (Worthington, 50-592-486). Dissociated cells were filtered through a 70um filter and spun down at 300 g for 10 min. Single-cell barcoding and complementary DNA (cDNA) library preparation were performed according to the previously published  protocol (Zilionus,R et al.,Nat protoc, 2017.) with following modification. A home-made microfluidic device was connected to the Drop-seq apparatus in the  Korea Brain Research Institute.  Amplified cDNA was not fragmented, and cDNA clean up and size selection was performed using SPRIselect beads (Beckman Coulter, B23317).  Single-cell libraries were sequenced on an Illumina NovaSeq 6000 system though a sequencer company (Macrogen).