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. 2014 Aug 12;9(8):e104513. doi: 10.1371/journal.pone.0104513

Figure 2. Specificity of YB-1 interactions with YB-1 mRNA 5′ UTR.

Figure 2

A. Streptavidin Sepharose-immobilized biotinylated RNA fragments (AβG mRNA, 100 nt nonspecific RNA fragment, 139 nt YB-1 mRNA 5′ UTR, 103 nt YB-1 mRNA 5′ UTR, and 136 nt YB-1 mRNA coding region fragment, 15 pmol each, were incubated with 300 µl of rabbit reticulocyte lysates. RNA-bound proteins were eluted, separated by SDS-PAGE, transferred onto a nitrocellulose membrane, and detected with anti-YB-1 antibodies. Lane 1 – rabbit reticulocyte lysate as a marker, lane 2 – control experiment without (w/o) RNA, lane 3 – AβG mRNA, lane 4–100 nt nonspecific RNA fragment, lane 5–139 nt YB-1 mRNA 5′ UTR, lane 6–103 nt YB-1 mRNA 5′ UTR, lane 7–136 nt YB-1 mRNA coding region fragment. B. YB-1-specific bands from lanes 3, 5, and 6 were quantified using ImageJ software, and RNA-bound YB-1 levels were normalized to the quantity of RNA used (in moles). The amount of 103 nt 5′ UTR-bound YB-1 was taken as 100%. Values are means of three independent experiments. Errors are two standard deviations.