DNA pol γ CRISPR Activation Plasmid (m): sc-422341-ACT

Polg encodes DNA polymerase gamma, the catalytic polymerase responsible for mitochondrial DNA replication and repair in mouse cells. DNA pol γ functions with accessory subunits to support mtDNA maintenance, linking oxidative phosphorylation capacity to mitochondrial biogenesis, nucleoid stability, and organelle stress responses. Disruption or altered activity in the POLG axis is associated with mtDNA depletion or accumulation of deletions, which can model mitochondrial dysfunction phenotypes relevant to neuromuscular and metabolic disease mechanisms. Polg is therefore widely used to interrogate mitochondrial genome integrity, replication fork dynamics, and retrograde signaling pathways that couple mitochondrial defects to nuclear transcriptional programs.

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

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

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