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Both the Tet-inducible vector pRMC2 (Addgene, plasmid 68940) 70 and the mKate insert were digested with KpnI-HF and EcoRI-HF (New England Biolabs), gel purified using a QIAquick PCR purification kit (Qiagen), and ligated using T4 DNA ligase (New England Biolabs) at 16 C for 16 h