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. 2008 Jun;14(6):1048–1056. doi: 10.1261/rna.942308

FIGURE 4.

FIGURE 4.

UV-cross-linking experiments suggest close proximity of the D3 bulge to ε-ε′ contact in D1. (A) D3 molecules containing a single 6-thio-dG substitution at A 615, A 599, or A 662 (highlighted by green circles) were obtained by enzymatic ligation of two chemically synthesized oligonucleotides at the site indicated by an arrow. (B) A schematic of the exD15 construct used in these studies. (C) Autoradiographs of representative sequencing gels showing specific cross-links of bulge nucleotides 599 and 662 to the residues forming the ε-ε′ contact in D1 (red circles). RT indicates lanes corresponding to the RT analysis of uncross-linked exD15 RNA (control for natural RT stops), X-RT analysis of the exD15 RNA after UV-cross-linking to D3 molecules containing single 6-thio-dG substitutions. Positions of 6-thio-dG substitutions are shown on top. Data represent the results of at least five independent experiments. A figure showing a primary cross-linking gel with indicated cross-linking efficiencies and a larger portion of the sequencing gel is available as supplementary material upon request (please send an e-mail message to anna.pyle@yale.edu).