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. Author manuscript; available in PMC: 2018 Mar 20.
Published in final edited form as: Methods Enzymol. 2017 May 5;591:119–157. doi: 10.1016/bs.mie.2017.03.003

Figure 2.

Figure 2

Western blot to quantify expression of exogenous WT and variant protein expression. A. The level of exogenous protein expression above endogenous protein expression is determined using a primary antibody against the BER enzyme of interest. The signal produced in lysates from cells not cultured with doxycycline (exogenous + endogenous protein) is divided by the signal produced in lysates from cells cultured with doxycycline (endogenous protein only) to calculate the fold increase of protein expression as a result of exogenous protein expression. To properly characterize the cellular effects of a human somatic or germline BER variant, it is important to maintain expression levels of the exogenous protein that are close to the levels of the endogenous protein. B. Expression of WT and variant is detected using a primary antibody against the HA epitope tag. Expression is normalized by dividing the HA signal by the tubulin signal.