GSM6720072: scRNA-seq of M1; Rattus norvegicus; RNA-Seq
Mandible were dissected under a stereomicroscope. The tooth germs of first mandibular molars and second mandibular molars were carefully isolated respectively, and then cut into small pieces. Subsequently, the small pieces were digested in 3 mg/mL Collagenase type I (Sigma-Aldrich) and 3 mg/mL Dispase II (Sigma-Aldrich) and 20U/ml Dnase I (Solarbio) at 37 ℃ for 1h, and then filtered through a 40 μm cell strainer (BD Biosciences). After washed with cold PBS, the cells were resuspended in PBS containing 0.04% BSA. 10x Genomics Chromium platform was used to capture and barcode cells to generate single-cell Gel Beads-in-Emulsion (GEMs) by following the manufacturer's protocol. Briefly, along with the reverse transcription master mix, cell suspensions of all samples were loaded onto the same 10x Genomics Single Cell 3′ Chip. During this step, cells were partitioned into GEMs along with gel beads coated with oligonucleotides. These oligonucleotides enable mRNA capture by 30 bp oligo-dT and provide barcodes to index cells (16 bp) as well as transcripts (12 bp unique molecular identifier; UMI). Following reverse transcription, cDNAs with both barcodes were amplified, and a library was constructed using the Single Cell 3′ Reagent Kit (v3 chemistry) for each sample. The resulting libraries were sequenced on an Illumina NovaSeq 6000 System.