Fig. 1

NOS1 is essential for cell proliferation and tumor growth of melanoma. A Schematic diagram of NOS1-targeting guide RNA (gRNA) sequences. B Knockout of NOS1 gene was confirmed by PCR assay with 131bp compare to 318bp in wide-type allele. A2, A4, A5 and A6 clone displayed NOS1 deletion. C Western blot showed that NOS1 knockout in A5 clone. D The sequence analysis confirmed A5 clone with 180bp deletion and 1bp mutation. E MTT assay showed NOS1-KO cells grow slower than WT cells (***\(\hbox {p}<0.001\)). F Plate formation assay indicated the clone number of NOS1-KO group was less than WT group. (*\(\hbox {p} < 0.05\)). G The reprehensive image (left panel ) of EdU staining(red) for proliferation cells of NOS1-KO and WT cells , and the proliferation cells was decreased in NOS1-KO cells compared to WT cells (right panel) (Scale bars, \(500 \; \upmu \hbox {m}\). ***\(\hbox {p} < 0.001\)). H Flow cytometry assay (PI staining) showed cell cycle was blocked at G1 phase by NOS1-KO. I The tumors of NOS1-KO and WT A375 in BALB/c-nu mouse at the termination of experiment (n = 5, left panel). The tumor weight (middle panel) and tumor growth curves (right panel) in NOS1-KO group were lower than WT group (*\(\hbox {p} < 0.05\))