Cloning and Knockdown of ILF3 Gene

The complementary DNA encoding ILF3 was PCR-amplified by the Ex Taq DNA polymerase hot-start version (Takara) and subcloned into the Nhe I and Kpn I sites of pcDNA3.1(+) plasmid (Thermo Fisher Scientific). The sequences of primers were as follows: 5′-CTAGCTAGCGATAAGCAAAAGTTTGATTTCCAG-3′ (sense) and 5′-GGGGTACCGGAGTAAGTGCAGAAGGTAGA-3′ (anti-sense). The empty plasmid pcDNA3.1(+) was used as negative control. To knock down ILF3 expression, two oligonucleotides for shRNAs were synthesized and inserted into the shRNA expression vector pGPH1/Neo (GenePharma, Shanghai, China). The sequences of shRNAs were 5′-ccaaggaacTcTaTcacaa-3′ for sh-ILF3-122 (link) and 5′-CCACTGATGCTATTGGGCATCTAGA-3′ for sh-ILF3-2.19 (link) A scrambled nontargeted shRNA was used as a negative control. The transfections of plasmids were carried out using Lipofectamine 3000 (Thermo Fisher Scientific) according to the protocol.