title: Illumina HiSeq 2500 paired end sequencing; RNA-seq of Mus musculus primary neural stem cells with MHC I knockout
design: RNA-seq of Mus musculus primary neural stem cells with MHC I knockout
library_construction_protocol: Hippocampi were dissected from postnatal day 1 (P1) C57/BL6, KD-/-, K-/-, or D-/- mice and mechanically dissociated by fine mincing and enzymatically digested for 30 minutes at 37ºC in DMEM media containing 2.5U/ml Papain, 1U/ml Dispase II , and 250U/ml DNase I. NSCs were purified using a 65% Percoll gradient and cultured (Neural Basal A medium supplemented with 2% B27 without Vitamin A, 1% Glutamax, 1% Penicillin Streptomycin, 10ng/ml EGF, 10ng/ml bFGF) as a monolayer on poly-D-lysine and laminin-coated plates at a density of 105 cells/cm2.  mRNA of NSCs was isolated by lysis with TRIZOL Reagent (Thermo Fisher Scientific), separation with chloroform (0.2 mL per mL TRIZOL), and precipitated with isopropyl alcohol.   Libraries were prepared using the Illumina Nextera XT DNA Sample Preparation Kit according to the manufacturers protocol. cDNA was validated using KAPA qPCR-based library quantification kit.
attributes.tag: Experimental Factor: genotype
attributes.value: H2-K1 knockout