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. 2015 Jun 4;96(6):1001–1007. doi: 10.1016/j.ajhg.2015.04.022

Figure 3.

Figure 3

Cell Survival Curves after MMC or UV Treatment

(A) MMC sensitivity assay. AP65P-hTERT cells were exposed to varying concentrations of MMC for 24 hr, washed, and seeded into 6-well plates. After 5–7 days, surviving cells were counted using a LUNA digital automated cell counter (Logos Biosystems) or stained with 0.006% crystal violet/25% methanol solution. GM6914 (FA-A) and its complemented cells were included as control.

(B) Immunoblotting using the indicated antibodies (listed in Table S3). AP65P-hTERT cells were transfected with control (siLuc) or siRNAs targeting UBE2T (siUBE2T#1 and #2) (final 10 nM), seeded, and treated with MMC for 24 hr or UV irradiated. siRNAs (sequence listed in Table S2) were synthesized by Life Technologies and transfected using Lipofectamine RNAiMAX.

(C) MMC or UV sensitivity in AP65P-hTERT cells depleted of mutated UBE2T protein.

(D) UV sensitivity in AP65P-hTERT cells with or without UBE2T lentiviral transduction. The sensitivity assays were repeated at least three times, and a representative dataset with mean and SD of the triplicate cultures is shown.