TBX4 CRISPR Activation Plasmid (h): sc-405242-ACT

TBX4 encodes a T-box family transcription factor that binds DNA through a conserved T-box domain to regulate developmental gene expression programs. In human biology, TBX4 is a key driver of hindlimb morphogenesis and mesenchymal differentiation, coordinating transcriptional networks that intersect with FGF, WNT, BMP, and SHH signaling during patterning and tissue specification. Altered TBX4 dosage or regulatory activity has been linked to congenital limb malformations and pediatric pulmonary vascular disease, reflecting its role in developmental and organogenesis pathways. As a nuclear regulator, TBX4 is frequently studied for its effects on lineage commitment, transcriptional circuitry, and enhancer-promoter control.

TBX4 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous TBX4 expression without altering the underlying DNA sequence.

TBX4 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the TBX4 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 TBX4 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous TBX4 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native TBX4 locus and enabling the study of TBX4-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of TBX4 pathway restoration in tumor cells with silenced or reduced TBX4 expression.

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