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. 2019 Jul 15;11:6615–6624. doi: 10.2147/CMAR.S208708

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© 2019 Gong et al.

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miR-625 directly targets ALDH1A1. (A) The schematic representation of complimentary sequences between miR-625 and 3ʹ-UTR of ALDH1A1. (B) SGC7901 cells were transfected with antagomir-625 or antagomir-NC in combination with luciferase reporter construct containing wild-type or mutant 3ʹ-UTR of ALDH1A1. At 2 days after transfection, the luciferase activity was measured. The firefly luciferase activity was normalized to that of Renilla luciferase. Each treatment condition was performed in triplicate. (C) SGC7901 cells were transfected with mimic-mir625 or mimic-NC in combination with luciferase reporter construct containing wild-type or mutant 3ʹ-UTR of ALDH1A1. At 2 days after transfection, the luciferase activity was measured. The firefly luciferase activity was normalized to that of Renilla luciferase. Each treatment condition was performed in triplicate. (D) The mRNA level of ALDH1A1 in SGC7901/ADR and SGC7901/VCR cells was determined by qRT-PCR analysis. Results were normalized to U6 snRNA. The expression relative to that in parental SGC7901 cells is shown. Each column represents the mean value from 3 replicates. (E) The protein level of ALDH1A1 in parental SGC7901 cells, SGC7901/ADR cells and SGC7901/VCR was determined by Western blot analysis. β-actin was utilized as a loading control. The representative results from 3 independent experiments are shown. (F) SGC7901 cells were transfected with antagomir-625 or antagomir-NC. At 2 days after transfection, the protein level of ALDH1A1 was determined by Western blot analysis. (G) SGC7901/ADR cells and SGC7901/VCR cells were transfected with mimic-mir625 or mimic-NC. At 2 days after transfection, the protein level of ALDH1A1 was determined by Western blot analysis. β-actin was utilized as a loading control. The representative results from 3 independent experiments are shown. Data are presented as the mean ± SEM. Two-tailed Student’s t-test. **P<0.01; NS, not significant.