NextSeq 500 paired end sequencing; Single-cell RNA-seq (10x Chromium V3) of heterogeneous spheres consisting of melanoma cell line and two types of human primary dermal fibroblasts
Heterogeneous spheres were prepared from primary human fibroblasts and G361 melanoma cell line (commercially available; CVCL_1220). Fibroblasts were isolated from photo-aged skin of an adult and from juvenile photo-protective skin of child. Heterogeneous spheres were washed in PBS and digested in 0.25% trypsin + EDTA solution (1:1). Cell suspensions were inoculated in DMEM + 10% fetal bovine serum for 72 hours in hanging drops. Heterogeneous spheres were cultured for additional 24 hours in non-adhesive culture dishes.  Immediately after tissue dissociation, single cell libraries were prepared on the 10x Chromium with a target of 4,000 cells. Chromium Single Cell 3สน GEM, Library & Gel Bead Kit v3, 4 rxns PN-1000092 was used. Immediately after tissue dissociation, single cell libraries were prepared on the 10x Chromium with a target of 4,000 cells. Chromium Single Cell 3สน GEM, Library & Gel Bead Kit v3, 4 rxns PN-1000092 was used.