We next addressed whether Mcm10 enables CMG to bypass the lagging strand streptavidin or whether the apparent stronger helicase activity of CMG helicase provided by Mcm10 enables CMG to displace the lagging strand blocks (Figure 6). To determine if the streptavidin was displaced or retained on DNA during CMG unwinding we 32P-labeled the biotinylated strand, which is gel shifted by streptavidin, and repeated the assay in the presence of excess biotin which traps displaced streptavidin and prevents it from rebinding biotinylated DNA. The control reactions, adding the biotin trap before streptavidin, show >96% inhibition of the streptavidin gel shift of the biotinylated DNAs (Figure 6—figure supplement 1). Upon performing the experiment using dual biotinylated-streptavidin blocks on the lagging strand template, the results show that nearly all of the unwound product DNA retains the streptavidin (i.e. it is gel shifted) in the presence of the excess biotin trap. Hence, CMG-Mcm10 bypasses lagging strand streptavidin blocks without displacing them from DNA.