ExtFig3E_RACE_INSIG1_SLC7A5_R2_LG306_annotated.tiff

HEK293T ishXrn1 cells were treated with doxycycline for 3-4 days to induce knock down of Xrn1, then transfected with the luciferase reporters containing 99 bp insertions from the BCAP1, INSIG1, SLC7A5, STOML2, YKT6, and TUBA1B genes, and where indicated, with PR8 PA-X. RNA was extracted and used to run 5’ RACE with primers annealing to luciferase sequences. DNA bands were purified and sequenced to confirm their identities

将HEK293T ishXrn1细胞经强力霉素（doxycycline）处理3至4天以诱导Xrn1基因敲低，随后转染携带来自BCAP1、INSIG1、SLC7A5、STOML2、YKT6及TUBA1B基因的99 bp插入片段的荧光素酶报告基因质粒；若实验有标注，则同时共转染PR8 PA-X。提取RNA后，使用靶向荧光素酶序列的引物开展5' 快速扩增cDNA末端（5' Rapid Amplification of cDNA Ends, 5' RACE）实验。对扩增得到的DNA条带进行纯化并测序，以确认其序列身份。