biostudies-arrayexpressNaiqi WangMus musculushttps://www.ebi.ac.uk/biostudies/studies/E-MTAB-16518Weaning mice were fed with SF-NIH31 or SF-AIN93G diets for four weeks, followed by implantation of B16-OVA melanoma cells. CD45+ leukocytes cells were sorted from tumor on 15 days post implantation, followed by single cell RNA sequencing or TCR sequencing.biostudies-arrayexpressLibrary Construction - For gene expression library construction, 50 ng of amplified cDNA was fragmented, end-repaired, adaptor ligated, and subjected to double-sided size selection using SPRIselect beads. Libraries were then indexed via PCR (98 °C for 3 min; 13 cycles of 98 °C for 20 s, 54 °C for 30 s, 72 °C for 20 s; final extension at 72 °C for 5 min), followed by a second double-sided SPRIselect cleanup. Library quality was assessed using an Agilent TapeStation, and sequencing was performed on a DNBSEQ-G400 platform using following configuration: 29 bp Read1, 8 bp N5 Index, 8 bp N7 Index, 90 bp Read2. For scTCR libraries, 3 μL of cDNA post amplification and purification was used for targeted TCR enrichment. The first enrichment was performed using mouse TCR primers 1, followed by SPRIselect bead cleanup. A second enrichment step with mouse TCR primers 2 was conducted, followed by double-sided size selection using SPRIselect beads. Library quality was assessed using an Agilent TapeStation. Enriched products underwent fragmentation, end-repair, adapter ligation, and further SPRIselect bead purification. Libraries were then indexed with adapter-specific primers, and final quality control was performed using an Agilent TapeStation.Sample Collection - Mice were fed with SF-AIN93G(SpecialtyFeed) or SF-NIH31 (SpecialtyFeed) for four weeks, followed by subcultaneous injection of 1*10^5 B16-OVA cells. At day 18 post implantation, CD45+ cells were sorted from processed tumours using FACS.Nucleic Acid Extraction - scRNA-seq and scTCR-seq libraries were prepared using the SeekOne® DD Single Cell 5′ Transcriptome-seq Kit or Single Cell BCR/TCR Enrichment Kit for mouse T cells (v1.3 Chemistry), following the manufacturer’s protocol. Briefly, FACS-sorted cells were washed twice with PBS containing 2% FBS, and approximately 20,000 cells were loaded into the SeekOne® Digital Droplet System for single-cell encapsulation and barcoding. After droplet breakage, cDNA was purified using magnetic cleanup beads and amplified by PCR (98 °C for 3 min; 13 cycles of 98 °C for 20 s, 63 °C for 30 s, 72 °C for 1 min; final extension at 72 °C for 5 min).Sequencing - Sequencing was carried out on a DNBSEQ-G400 platform with the following read configuration: 29 bp Read 1, 8 bp N5 Index, 8 bp N7 Index, and 150 bp Read 2.OrganizationMINSEQE ScoreAssays and DataMAGE-TAB FilesSequence Alignment - scRNA-seq reads were aligned to the mm10 reference genome and quantified using seeksoultools rna (seekgene, v.1.2.0). Filtered gene-barcode matrices, containing only barcodes with unique molecular identifier counts that passed the cell detection threshold, were used for downstream analysis. scTCR reads were aligned to the mm10 reference genome, and consensus TCR annotation was performed using seeksoultools vdj (seekgene, v.1.2.0). TCR annotation followed the 10x seeksoultools vdj pipeline.UnknownTranscriptomicsGenomicsProteomicsBD FACSAria III cell sorter (BD Biosciences)DNBSEQ-G400SeekOne® Digital Droplet System (Seek Gene)S1000 Thermal Cycler (BioRad)RNA-seq of coding RNA from single cellsMus musculusNaiqi WangfalseSingle cell RNA and TCR sequencing of leukocytes in melanoma tumor and draining lymph node (dLN) from mice fed with SF-NIH31 or SF-AIN93G dietWeaning mice were fed with SF-NIH31 or SF-AIN93G diets for four weeks, followed by implantation of B16-OVA melanoma cells. CD45+ leukocytes cells were sorted from tumor on 15 days post implantation, followed by single cell RNA sequencing or TCR sequencing.2026-05-01T00:00:00Z2026-05-01T01:03:27.137Z2026-01-13T16:46:19.638ZE-MTAB-16518ERP187550E-MTAB-16107EFO_0002944EFO_0004170EFO_0005684EFO_0004917EFO_0005518EFO_0004184