Nocturnin CRISPR Activation Plasmid (h): sc-405350-ACT

Human NOCT encodes Nocturnin, a circadian-regulated CCR4-like deadenylase that shortens poly(A) tails to control mRNA stability and translational output. Nocturnin helps couple clock circuitry to metabolic gene programs, influencing pathways linked to lipid handling, energy balance, and nutrient-responsive transcription. Through post-transcriptional remodeling of transcripts, NOCT contributes to rhythmic regulation of cellular homeostasis and stress adaptation. Dysregulated NOCT expression or activity has been associated with altered metabolic phenotypes and has been explored in the context of inflammation and cancer-related gene expression programs.

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

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

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