cyclin E CRISPR Activation Plasmid (h): sc-400105-ACT

Human CCNE1 encodes cyclin E, a core regulator of the G1/S transition that forms an active kinase complex with CDK2 to promote DNA replication origin licensing and S-phase entry. Cyclin E–CDK2 signaling integrates mitogenic inputs with checkpoint control by coordinating phosphorylation of substrates involved in E2F-dependent transcription, replication timing, and centrosome duplication. Dysregulated CCNE1 expression perturbs cell-cycle fidelity and genome stability, linking cyclin E activity to replication stress, chromosomal instability, and oncogenic cell proliferation. Consequently, CCNE1 is widely studied in pathways governing cell-cycle progression, DNA damage responses, and mechanisms of cell growth control in human model systems.

cyclin E CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous CCNE1 expression without altering the underlying DNA sequence.

cyclin E CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the CCNE1 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 CCNE1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous cyclin E expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native CCNE1 locus and enabling the study of cyclin E-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of cyclin E pathway restoration in tumor cells with silenced or reduced CCNE1 expression.

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