Next Generation Sequencing Facilitates Quantitative Analysis of C57BL/6J mice Incisor Transcriptomes at single cell level.

Purpose: Analysis the heterogeneity of the MSC population in the adult mouse incisor, using single cell RNA-sequence. Results: About 9,318 cells (~63,000 reads per cells) were successfully barcoded and their transcriptomes sequenced. Overall design: Methods: Mandibles were carefully dissected under a stereomicroscope. The surrounding jaw bones were removed by scalpel and the proximal region was then severed. Dental pulp with dental epithelium was isolated, cut into pieces, collected in cold PBS, and digested with 2mg/ml collagenase type I dissolved in a-MEM for 30 min at 37 ?. After incubation, the suspension was homogenized by pipetting and digestion was terminated by adding 10% FBS. The digested tissues were centrifuged at 300g for 5 min and resuspended in 10% FBS. Twenty thousand cells were loaded into the 10X Chromium system with targeted cell recovery of 10,000 cells to be barcoded for scRNA-seq using a Single Cell 3' Library Kit v3. Sequencing was performed on the Illumina Novaseq System.