GSM8727834: Popliteal Lymph Node, VACV-gB injection, Spatial transcriptomics, 21dpi; Mus musculus; OTHER
Slide-seq Spatial Transcriptomics libraries for all tissue sections were prepared using the Curio Seeker Spatial Transcriptomics kit (Curio Biosciences), following the manufacturer's instructions. Briefly, fresh-frozen tissue was mounted on 3mm x 3mm Curio Seeker tiles, followed by RNA hybridization, reverse transcription, enzymatic digestion, and bead detachment from the glass tiles. This was followed by second strand cDNA synthesis and amplification. The cDNA library from each sample was divided into two portions. For spatial transcriptomics, the cDNA ibraries were prepared with the Nextera XT DNA sample preparation kit following Curio instructions. The samples were then pooled and sequenced on an Illumina NextSeq 2K system with a P2 flow cell and a 100-cycle kit, with read configurations as follows: Read 1 = 50 bp, Read 2 = 72 bp, Index 1 = 8 bp, and Index 2 = 8 bp. For the Long read, the cDNA library was processed with hybridization-based target enrichment of TCR and BCR transcripts and rolling circle amplification to concatemeric consensus (R2C2). The processed libraries were prepared using Ligation Sequencing Kit V14 from the Oxford Nanopore before sequencing by MinION (R10.4.1) flowcell for at least 48 hours.