Single-cell RNA-seq of Hepa1-6 Tumor Cells

Hepa1-6 tumors were treated with PBS, Virus@gel or V-Navo@gel (1 × 10⁷ PFU) intratumorally for 12 days. Then the tumors were dissociated and stored in the sCelLive^TM Tissue Preservation Solution (Singleron Bio Com, Nanjing, China) on ice after the surgery within 30 min. Tumors were digested with 2 mL sCelLive^TM Tissue Dissociation Solution (Singleron) by Singleron PythoN® Automated Tissue Dissociation System (Singleron) at 37 °C for 15 min. The solution was then centrifuged at 500 g for 5 mins and suspended softly with PBS. Then the single-cell suspensions (1 × 10⁵ cells/mL) were loaded into microfluidic devices using the Singleron Matrix® Single Cell Processing System (Singleron). Subsequently, the scRNA-seq libraries were constructed according to the protocol of the GEXSCOPE® Single Cell RNA Library Kits (Singleron)^{46 (link)}. Individual libraries were diluted to 4 nM and pooled for sequencing. At last, pools were sequenced on Illumina novaseq6000 with 150 bp paired end reads.