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. 2011 Mar 11;39(12):5245–5254. doi: 10.1093/nar/gkr132

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© The Author(s) 2011. Published by Oxford University Press.

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.5), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 1. — Reconstitution of the XBP1u mRNA splicing in vitro. (A) Splicing assay. The reaction mixtures (40 µl) contained recombinant human IRE1α (rIRE1α), transcribed XBP1u RNA (XBP1u) and rabbit erythrocyte lysate (REL). After the reaction, the relative ligase activity was examined as described in ‘Materials and Methods’ section. The amounts of REL added to the reactions are: 0 µg (lane 1), 25 µg (lane 2), 125 µg (lane 3), 250 µg (lane 4), 500 µg (lane 5). The positions of the RT-PCR products representing the XBP1u (U) and XBP1s (S) RNAs are indicated. (B) Determination of the nucleotide sequence of the band that corresponds to the XBP1u (U) and XBP1s (S) RNAs in (A). The sequence around the splice junction revealed that two halves of XBP1s RNA were joined at the correct position after the in vitro splicing reaction. Numbers in (B) denote nucleotide positions. The first A of the initiation methionine codon is set at 1.