In contrast to studies with pure CMG, studies of replication in Xenopus egg extracts show that two adjacent streptavidin blocks spaced 5 bp apart on the non-translocating (lagging) strand are quickly bypassed (within 10 min) by the replisome (Fu et al., 2011). The fact that our earlier study showed that isolated CMG is strongly inhibited by a dual streptavidin block, suggests that some other factor in the complete extract helps CMG bypass lagging strand blocks (Langston et al., 2017). To test whether Mcm10 may be the factor that facilitates CMG bypass of a lagging strand block we examined CMG unwinding of the forked DNA substrate with two biotinylated nucleotides on the duplex portion of the lagging strand template (see Figure 5A). Control reactions lacking Mcm10 show that CMG helicase is essentially shut down by the streptavidin blocks (Figure 5B, compare lanes 2–4 without streptavidin to lanes 5–7 with streptavidin) as previously observed (Langston and O'Donnell, 2017). But surprisingly, when Mcm10 is added, the rate and amount of unwinding are the same in the presence and absence of streptavidin (Figure 5B, compare lanes 8–10 and 11–13). Hence, Mcm10 enables CMG to rapidly bypass or displace a dual streptavidin block on the lagging strand. This observation mirrors results observed in the Xenopus system, which was assumed (but not demonstrated) to bypass these blocks without displacement. Ability of CMG to bypass lagging strand blocks with Mcm10 present in the pure protein system resolves the discrepancies between use of purified CMG (without Mcm10), and observations in Xenopus extracts (Fu et al., 2011; Langston and O'Donnell, 2017). We revisit the issue of whether streptavidin blocks are bypassed or displaced later in this report.