ANKRD50 CRISPR Activation Plasmid (h): sc-412772-ACT

Human ANKRD50 encodes an ankyrin repeat–containing protein implicated in endosomal trafficking and membrane protein sorting, supporting the spatial control of signaling receptors and transporters. Reported functions link ANKRD50 to endosome-associated regulatory networks that influence receptor recycling, vesicular dynamics, and downstream pathway tuning, including processes that modulate growth-factor and immune signaling outputs. Through these roles, ANKRD50 is relevant to studies of cellular homeostasis, proteostasis, and signal transduction fidelity in differentiated and proliferative contexts. Altered endomembrane trafficking is a common feature across multiple disease mechanisms, making ANKRD50 a useful target for dissecting how trafficking perturbations reshape cellular signaling states.

ANKRD50 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous ANKRD50 expression without altering the underlying DNA sequence.

ANKRD50 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the ANKRD50 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 ANKRD50 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous ANKRD50 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native ANKRD50 locus and enabling the study of ANKRD50-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of ANKRD50 pathway restoration in tumor cells with silenced or reduced ANKRD50 expression.

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