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. 2018 Dec 7;15(12):1499–1511. doi: 10.1080/15476286.2018.1553481

Figure 5.

Figure 5.

LINC00673-mediated senescence is p53-dependent. (A) BrdU Cell Proliferation ELISA Kit was used to quantify cell proliferation 48 h after knockdown of p53, pRb and LINC00673 in A549 (n = 4). The reactions contained siPOOL negative control (12 nM) or a mix of 1 nM siPOOL p53, pRb and 10 nM siPOOL LINC00673 supplemented with siPOOL negative control for a final concentration of 12 nM. (B) BrdU incorporation in IMR-90 cells was measured as in A. The reactions contained siPOOL negative control (0.9 nM) or a mix of 0.3 nM siPOOL p53, pRb and LINC00673 supplemented with siPOOL negative control for a final concentration of 0.9 nM. (C) Relative RNA levels were determined by RT-qPCR at 48 h after knockdown in A549 cells (n = 3). The reactions contained siPOOL negative control (5 nM) or a mix of 1 nM siPOOL p53, pRb and 3 nM siPOOL LINC00673 supplemented with siPOOL negative control for a final concentration of 5 nM. GAPDH was used as reference gene. One representative Western Blot is shown (n = 3). (D) RT-qPCR as in C using IMR-90 cells (n = 4). The siPOOL concentrations were as described in B. One representative Western Blot is shown (n = 4). (E) SA-β-Gal accumulated in A549 cells 4 days after knockdown. The reactions contained siPOOL negative control (4 nM) or a mix of 3 nM siPOOL LINC00673 and 1 nM siPOOL p53 or pRb supplemented with siPOOL negative control for a final concentration of 4 nM (n = 4, scale bar = 100 µm). In A-F, the mean + SEM is shown. The statistical significance was determined per two-sided unpaired Student t test, with *, P < 0.05; **, P < 0.01; ***, P < 0.001.