Fig. 3

Knock out of LacRNA promotes breast cancer metastasis. a Schematic diagram of paired-gRNA plasmid construction. Paired gRNAs (pgRNAs) in one lentiviral backbone with two U6 promoters separately driving the two gRNAs. b Schematic diagram of the position of the four candidate sgRNA#1-#4 before and after LacRNA genome transcriptional start site (tss5) and the position of genomic PCR validation primers (L1 and R1 primers). c The knockout efficiency of LacRNA by CRISPR-Cas9 technology knockout in the MDA-MB-231 and BT549 cells. Values are expressed as the Mean ± SEM; n = 3 independent experiments. P values were determined by one way analysis of variance (ANOVA). ns, not significant, **P < 0.01, ***P < 0.001. d After infection with paired-gRNA, puromycin was screened for 5 and 10 days, and the deletion of genomic DNA was verified by PCR. e, f Transwell migration and invasion assays of BT-549 and MDA-MB-231 cells with LacRNA knockout by CRISPR/Cas9 technology. The stable cells established via the CRISPR/Cas9 knockout technology were designated as KO. Values are expressed as the Mean ± SEM in c, e and f. n = 3 independent experiments. P values were determined by one-way analysis of variance (ANOVA). ns, not significant, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001