T-bet/TBX21 CRISPR Activation Plasmid (h): sc-400480-ACT

TBX21 encodes T-bet, a T-box transcription factor that functions as a master regulator of type 1 immune differentiation. T-bet integrates cytokine and TCR signaling to shape chromatin accessibility and transcriptional programs controlling IFNG and other Th1-associated genes, influencing NK cell maturation and cytotoxic effector function. Through cross-regulatory interactions with lineage-defining factors, TBX21 helps balance Th1 versus Th2/Th17 polarization and coordinates responses downstream of IL-12/STAT4 and IFN/STAT1 pathways. Dysregulated TBX21 activity has been associated with immune-mediated inflammation, susceptibility to infection, and altered tumor immune surveillance, making it a valuable node for mechanistic studies of immunopathology.

T-bet/TBX21 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous TBX21 expression without altering the underlying DNA sequence.

T-bet/TBX21 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the TBX21 locus in human cell lines. The system is built around a catalytically inactive Cas9 (dCas9) carrying two inactivating mutations (D10A and N863A) that eliminate nuclease activity while preserving DNA binding. This dCas9 is fused to VP64, a potent transcriptional activator, and is co-expressed with a blasticidin resistance gene for selection. The second plasmid encodes the MS2-p65-HSF1 fusion protein, a secondary activator complex that works in concert with dCas9-VP64, alongside a hygromycin resistance gene. The third plasmid encodes a target-specific 20 nt sgRNA fused to two MS2 RNA aptamers that recruit the MS2-p65-HSF1 complex to the activation site, accompanied by a puromycin resistance gene. The three plasmids are delivered at a 1:1:1 mass ratio for balanced expression of all system components.

Once assembled at the target locus, the SAM complex binds within approximately 200 bp upstream of the TBX21 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous T-bet/TBX21 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native TBX21 locus and enabling the study of T-bet/TBX21-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of T-bet/TBX21 pathway restoration in tumor cells with silenced or reduced TBX21 expression.

For Research Use Only. Not Intended for Diagnostic or Therapeutic Use.