Ninein CRISPR Activation Plasmid (h): sc-402270-ACT

Human NIN encodes ninein, a centrosomal coiled-coil protein that localizes to the mother centriole and pericentriolar material, where it anchors microtubule minus ends and helps organize non-centrosomal microtubule arrays. Ninein contributes to centrosome maturation, spindle assembly, ciliogenesis, and maintenance of cell polarity by coordinating microtubule nucleation and anchoring dynamics during the cell cycle. Through these roles, NIN links centrosome integrity with mitotic fidelity and cytoskeletal architecture, processes frequently perturbed in proliferative and neurodevelopmental disorders. Altered ninein function has been studied in the context of defects in ciliary signaling, cell migration, and chromosomal instability phenotypes relevant to disease mechanism research.

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

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

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