DNA pol δ 4 CRISPR Activation Plasmid (h2): sc-408268-ACT-2

Human POLD4 encodes DNA polymerase delta subunit 4 (Pol δ4), an accessory component of the Pol δ holoenzyme that supports high-fidelity lagging-strand DNA synthesis during replication and contributes to DNA repair synthesis. As part of the replication machinery, POLD4 functions within core pathways governing replication fork progression, Okazaki fragment processing, and genome stability checkpoints, coordinating with PCNA-dependent polymerase activity. Altered Pol δ subunit balance or replication stress involving POLD4 has been linked to increased mutational burden and chromosomal instability, features frequently observed in cancer and other genome maintenance disorders. POLD4-targeted gene editing and functional assays enable studies of polymerase complex assembly, replication/repair pathway choice, and the cellular consequences of perturbed DNA synthesis fidelity in human model systems.

DNA pol δ 4 CRISPR Activation Plasmid (h2) provides a targeted, non-destructive approach to upregulating endogenous POLD4 expression without altering the underlying DNA sequence.

DNA pol δ 4 CRISPR Activation Plasmid (h2) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the POLD4 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 POLD4 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous DNA pol δ 4 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native POLD4 locus and enabling the study of DNA pol δ 4-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of DNA pol δ 4 pathway restoration in tumor cells with silenced or reduced POLD4 expression.

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