GSM8666017: Cochlea, 10 weeks, Bio rep 5; Mus musculus; RNA-Seq
Whole cochlear and vestibular organs were microdissected from the inner ears of 10-weeks-old CBA/J mice in petri dishes containing cold L-15 media (Gibco). The sensory epithelia were subjected to enzymatic digestion by transferring into 1.5 ml eppendorf tubes containing 1 ml/mg collagenase IV (Sigma) in L-15 medium for 10 minutes at room temperature. Then the cells were resuspended in 400 µL of enzyme-free DMEM containing 10% FBS and single-cell suspensions were obtained by trituration using a trimmed 200 ul pipette tip followed by filtering with a 40 uM strainer. The cells were pelleted by centrifuging at 300 xg for 5 mins at 4°C, then resuspended in media and subsequently quantified the number of cells. 7-8 mice were used for each biological replicate, and 6 and 4 biological replicates were obtained for cochlea and vestibule respectively. Single-cell capture and library preparation were performed using droplet-based microfluidic technology, following the manufacturer's instructions for the Chromium Next GEM Single Cell 3ʹ v3.1 dual index kit. The quality of the obtained libraries was assessed via high-sensitivity DNA kits (Agilent Technologies) using Agilent 2100 Bioanalyzer prior to sequencing with Illumina NovaSeq 6000.