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RNA was isolated from cells using the RNeasy Mini kit (Qiagen, ♯74104) and Xbp1 bands (both spliced and unspliced) from MEFs were amplified directly with primers (Fwd-5′GAAGAGAACCACAAACTCCA 3′; Rev-5′GGATATCAGACTCAGAATCT 3′) using OneStep RT-PCR Kit (Qiagen, ♯210210). Fragments were analysed by gel electrophoresis using 2.5% agarose gel in TAE buffer. 26 base pairs are excised out from unspliced Xbp1 to yield the spliced fragment.
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