(A) Illustration of the helicase reaction scheme. See Materials and methods for details. (B) Unwinding assays on dual biotinylated forks with or without streptavidin and with or without Mcm10, as indicated. (C) Same as in panel B except reactions contained or omitted MTC complex instead of Mcm10. (D) Illustration of the replisome reaction scheme. Reactions contained a synthetic minifork with 32P-5’ end-labeled primer and a dual biotin-streptavidin (SA) block on the lagging strand template. CMG and ATP were added for 3 min to allow CMG to bind DNA and translocate to the SA blocks, then Pol ε, RFC and PCNA were added (±MTC) along with all four dNTPs to assemble and extend the primer up to the blocked CMG, followed by addition of Mcm10 (or no protein). See Materials and methods for details. (E) Timed aliquots after ± Mcm10 addition were analyzed in a 10% urea PAGE gel. The primer, two stalled products and the full-length product are indicated to the right of the gel. Also see Figure 5—figure supplement 1.