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. 2021 Sep 8;10:e72286. doi: 10.7554/eLife.72286

Figure 2. Both co-directional (CD) and head-on (HO) R-loops perturb normal fork progression.

(A) Schematic illustrating expected sizes of replication products. (B) Denaturing agarose gel analysis of replication products obtained on R-loop-free template. Left lead: Leftward leading strands; Right lead: Rightward leading strands. (C) Native (top) and denaturing (bottom) agarose gel analyses of replication products obtained on templates harboring Airn sequence in CD or HO orientation. Stall: Stalled rightward leading strands; Restart: Rightward leading strand restart product; Full-length: full-length rightward leading strand; RI: replication intermediates. (D) Schematic illustrating replication products observed in (C). (E) Two-dimensional gel analysis of replication products obtained in presence of R-loops (corresponding to lanes 4 and 8 in (C)). Products were digested with ClaI. (F) Replication products obtained in the absence or presence of RFC/PCNA or Pol δ.

Figure 2—source data 1. Both co-directional (CD) and head-on (HO) R-loops perturb normal fork progression.
Figure 2—source data 2. Both co-directional (CD) and head-on (HO) R-loops perturb normal fork progression.
Figure 2—source data 3. Both co-directional (CD) and head-on (HO) R-loops perturb normal fork progression.

Figure 2.

Figure 2—figure supplement 1. Both co-directional (CD) and head-on (HO) R-loops perturb normal fork progression.

Figure 2—figure supplement 1.

(A) Replication products obtained on CD and HO templates with and without ClaI digestion post replication. (B) Conventional time course analyses of replication reactions on CD and HO templates. Time indicates minutes after addition of Mcm10 (origin firing).
Figure 2—figure supplement 1—source data 1. Characterization of replisome encounters with co-directional (CD) and head-on (HO) R-loops.
Figure 2—figure supplement 1—source data 2. Characterization of replisome encounters with co-directional (CD) and head-on (HO) R-loops.
Figure 2—figure supplement 2. Characterization of replisome encounters with co-directional (CD) and head-on (HO) R-loops.

Figure 2—figure supplement 2.

(A) Top: Schematic of products expected after digestion of full-length or restart replication products with Nt.BbvCI or Nb.BbvCI. Bottom: Denaturing gel analysis of replication products digested as indicated. (B) Replication products obtained on CD and HO templates in the absence or presence of Csm3-Tof1 and Mrc1 (CTM).
Figure 2—figure supplement 2—source data 1. Characterization of replisome encounters with co-directional (CD) and head-on (HO) R-loops.