MCM2 CRISPR Activation Plasmid (m): sc-421591-ACT

Mcm2 encodes the MCM2 subunit of the minichromosome maintenance (MCM2–7) helicase complex, an essential component of the eukaryotic DNA replication licensing machinery. MCM2 participates in pre-replication complex assembly at replication origins and supports S-phase progression by enabling origin firing and replication fork unwinding in coordination with CDC45 and the GINS complex. Tight regulation of MCM2 abundance and chromatin loading is linked to replication stress responses, ATR/CHK1 signaling, and maintenance of genome stability. Dysregulated MCM2 activity or expression is frequently associated with hyperproliferative states and chromosomal instability, making it a widely used marker and mechanistic node in studies of cell cycle control and DNA damage responses.

MCM2 CRISPR Activation Plasmid (m) provides a targeted, non-destructive approach to upregulating endogenous Mcm2 expression without altering the underlying DNA sequence.

MCM2 CRISPR Activation Plasmid (m) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the Mcm2 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 Mcm2 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous MCM2 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native Mcm2 locus and enabling the study of MCM2-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of MCM2 pathway restoration in tumor cells with silenced or reduced Mcm2 expression.

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