title: GSM9204060: NC-tumor-1; Mus musculus; RNA-Seq
library_construction_protocol: Tissues were cut into approximately 0.5-mm3 pieces in the RPMI-1640 medium (Invitrogen) with 1% Penicillin/Streptomycin, and enzymatically digested with the MACS Tumour Dissociation Kit _mouse (Miltenyi Biotec) at 37 °C for 30 min with agitation, according to the manufacturer's instructions. The dissociated cells were subsequently passed through a 70-μm and 40-µm cell-strainer (BD) and centrifuged at 300g for 10 min. After the supernatant was removed, the pelleted cells were suspended in red blood cell lysis buffer (Thermo Fisher) and incubated on ice for 2 min to lyse red blood cells. After washing twice with PBS (Invitrogen), the cell pellets were re-suspended in PBS (containing 0.04%BSA). DNBelab C Series High-throughput Single-Cell RNA Library Preparation Set (MGI, #940-001818-00 was utilized for scRNA-seq library preparation. In brief, the single-cell suspensions were converted to barcoded scRNA-seq libraries through steps including droplet encapsulation, emulsion breakage, mRNA captured beads collection, reverse transcription, cDNA amplification and purification. cDNA production was sheared to short fragments with 300-500 bp, and indexed sequencing libraries were constructed according to the manufacturer's protocol. Qualification was performed using Qubit ssDNA Assay Kit (Thermo Fisher Scientific) and Agilent Bioanalyzer 2100. All libraries were further sequenced by the DNBSEQ-T7 sequencing platform with pair-end sequencing. The sequencing reads contained 30-bp read 1 (including the 10-bp cell barcode 1, 10-bp cell barcode 2 and 10-bp unique molecular identifiers (UMI)), 100-bp read 2 for gene sequences and 10-bp barcodes read for sample index