Exogenous expression analysis of U1 snRNA mutation

To reveal the effect of U1 snRNA mutation, we performed exogenous expression analysis of U1 snRNA mutation. The pLKO.1-puro U6 sgRNA BfuAI stuffer lentiviral vector was modified by removing the internal U6 promoter. It was replaced by the U1 locus, including 393 bases of internal native U1 promoter, the U1 sequence, and 39 bases of 3’-flanking region. The r.3A>G mutation was introduced by site-directed mutagenesis. HEK-293T cells were transfected using Lipofectamine Plus with either pLKO.1-U1wt or pLKO.1-U1r.3a>g in duplicate. Messenger RNA library construction was performed based on oligo dT-based mRNA isolation using NEBNext® Poly(A) mRNA Magnetic Isolation Module. RNA Sequence was performed on NextSeq 550 using 100-bp paired-end mode. mRNA profiles of HEK293T with either U1 mutation or U1 wildtype vector.