Fig. 4

LINC00514 was a sponge for miR-28-5p. (a-b) Subcellular fractionation assay was used to determine the subcellular localization of LINC00514. a: BxPC-3 cells, b: SW1990 cells. (C) Sequence of WT-LINC00514, Mut-LINC00514 and miR-28-5p were conducted. (d-g) Luciferase reporter assay and RIP assay was performed to demonstrate that miR-28-5p was a downstream target of LINC00514. (h) Pull-down assay was conducted to detect the reaction between miR-28-5p and WT-LINC00514 or Mut-LINC00514. (i-j) Relative miR-18-5p expression level in BxPC-3 and SW1990 were determined by qRT-PCR. (K) The expression of miR-28-5p in PC tissue and normal tissue were detected by qRT-PCR. (L) Spearman’s rank correlation test was utilized to analyze the correlation between the levels of LINC00514 and miR-28-5p. (M-N) The miR-28-5p expression levels under LINC00514 silencing and LINC00514 overexpression were evaluated in vivo. (o-p) CCK-8 assay was performed to detect proliferation of cells transfected with LINC00514 shRNA and cells co-transfected with LINC00514 and miR-28-5p inhibitor. (q-r) Transwell migration and invasion assay were carried out to detect cell migration and invasion abilities. *p<.05, **p<.01, **p<.001. All experiments were repeated at least for three times and mean±SD was used to represent the final result. PC, pancreatic cancer; qRT-PCR, quantitative real-time polymerase chain reaction; RIP, RNA immunoprecipitation; SD, standard deviation; WT-lINC00514, wild type LINC00514; Mut-LINC00514, mutant LINC00514; CCK-8, cell counting kit‐8