GSM8101927: CnB1KO_S6 biol rep5; Mus musculus; RNA-Seq
For RNA-seq analysis, mouse brains were quickly dissected from normal slept (S6) or 6-h sleep-deprived (SD6) ABC-Ctrl (WT) and ABC-CnB1KO (KO) mice, rinsed with PBS, and flash frozen in liquid nitrogen (4 weeks after AAV-hSyn-Cre injection). After equilibration at -20 °C, mouse brains were sectioned at 200 μm on a cryostat microtome (Leica) under the protection of tissue OCT-Freeze medium. Locus coeruleus (LC) was further dissected from related slices and frozen in liquid nitrogen. Total RNA was isolated from LC (four to five biological replicates per group) using Trizol (ambion, 15596018) and precipitated by isopropanol added Sodium Acetate and Dr.GenTLE™ Precipitation Carrier (9094, Takara). Briefly, poly(A) mRNA was isolated from total RNA using the NEBNext Poly(A) mRNA Magnetic Isolation Module (E7490L, NEB). First-strand cDNA was synthesized with the SuperScript II Reverse Transcriptase (18064014, Invitrogen), and second-strand cDNA was synthesized using Second Strand cDNA Synthesis Kit (A48570, Invitrogen). Libraries were constructed with the NEBNext Ultra II DNA Library Prep Kit for Illumina (E7645L, NEB). Libraries were sequenced on Illumina HiSeq2500.