MISP CRISPR Activation Plasmid (h): sc-409029-ACT

Human MISP (mitotic spindle positioning) encodes an actin- and microtubule-associated protein that contributes to centrosome and spindle orientation, mitotic progression, and maintenance of epithelial architecture. MISP integrates cytoskeletal remodeling with cell-cycle control by coordinating cortical actin dynamics, focal adhesion-linked signaling, and proper bipolar spindle formation to support accurate chromosome segregation. Dysregulated spindle positioning and cytoskeletal organization are broadly associated with chromosomal instability and altered tissue polarity, making MISP a relevant target for studying mechanisms that couple adhesion, polarity, and division. In biomedical research, modulation of MISP expression is used to probe mitotic fidelity, cell shape control, and pathway dependencies in transformed and non-transformed human cell models.

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

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

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