Tom20 CRISPR Activation Plasmid (h): sc-400041-ACT

Human TOMM20 encodes Tom20, a core receptor of the translocase of the outer mitochondrial membrane (TOM) complex that recognizes N‑terminal targeting presequences and initiates import of nuclear‑encoded proteins into mitochondria. By controlling mitochondrial protein trafficking, Tom20 supports oxidative phosphorylation, mitochondrial dynamics, and proteostasis, and it influences cellular responses to energetic stress. Altered TOMM20 expression or function has been linked to mitochondrial dysfunction and perturbations in apoptosis and innate immune signaling, making it relevant to studies of cancer metabolism and neurodegenerative disease mechanisms. TOMM20 is therefore widely used as a mitochondrial marker and as a functional node connecting organelle biogenesis to cell survival pathways.

Tom20 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous TOMM20 expression without altering the underlying DNA sequence.

Tom20 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the TOMM20 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 TOMM20 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Tom20 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native TOMM20 locus and enabling the study of Tom20-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Tom20 pathway restoration in tumor cells with silenced or reduced TOMM20 expression.

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