DUSP1 and scramble shRNA were cloned into a pTRIPZ backbone. pTRIPZ plasmid and viral packaging plasmids were transfected into HEK 293 T cells with Lipofectamine Reagent (Thermo Fisher 18324012) (2 μg pMD2G, 3 μg psPAX2, and 5 μg pTRIPZ) and virus was collected and filtered after 48 h. Viral supernatant (4 mL) was supplemented with fresh media (2 mL) and Polybrene (8 μg/mL) and incubated with E006AA-hT cells for 4 h. After 48 h, cells were treated with puromycin (1 μg/mL). Once selected, shRNA expression was induced with 1 μg/mL doxycycline for 48 h.
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