SMC6 CRISPR Activation Plasmid (h): sc-402805-ACT

SMC6 encodes a core subunit of the SMC5/6 complex, an ATP-dependent chromosomal maintenance machine that safeguards genome stability during DNA replication and repair. SMC6 supports resolution of replication stress, repair of DNA double-strand breaks, and regulation of homologous recombination, contributing to chromosome segregation and telomere maintenance. Through these functions, SMC6 participates in DNA damage response and checkpoint-linked processes that constrain accumulation of chromosomal abnormalities. Dysregulation of SMC5/6 complex activity has been associated with genomic instability phenotypes relevant to cancer biology and inherited chromosome-breakage and replication-stress syndromes.

SMC6 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous SMC6 expression without altering the underlying DNA sequence.

SMC6 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the SMC6 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 SMC6 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous SMC6 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native SMC6 locus and enabling the study of SMC6-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of SMC6 pathway restoration in tumor cells with silenced or reduced SMC6 expression.

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