mRNA expression changes after SAM CRISPRa activator (MCP-p65AD-HSF1AD) expression in the primary effusion lymphoma cell line BC-3
CRISPR-based transcriptional activation (CRISPRa) has extensive research and clinical potential. In our related work, we showed that commonly used CRISPRa systems can exhibit pronounced cytotoxicity. Here, we investigated mRNA expression changes after the inducible expression of a fusion protein that is commonly used for CRISPRa, including the activation domains (ADs) of the transcription factors p65 and HSF1, components of the synergistic activation mediator (SAM) CRISPRa system. Overall design: BC3 cells that were stably transduced with the inducible lentiviral vector pTOZdeltaWRE-MPH or -hrGFP2 were plated at 400,000 cells/ml and induced with or without 100ng/ml doxycycline. 21 hour later, cells were recovered by centrifugation and subjected to dead cell removal (Miltenyi Biotec Dead Cell Removal Kit) as instructed. Resulting cell pellets were lysed in TRIzol (Invitrogen) and RNA was purified as instructed. MPH refers to a fusion protein of the MCP coat protein and the transactivation domains of p65 and HSF1.