myotubularin CRISPR Activation Plasmid (h): sc-403912-ACT

MTM1 encodes myotubularin, a lipid phosphatase that dephosphorylates phosphatidylinositol 3-phosphate and phosphatidylinositol (3,5)-bisphosphate, thereby regulating phosphoinositide signaling on endosomal and membrane trafficking compartments. Through control of endocytosis, autophagy-related membrane dynamics, and cytoskeletal organization, myotubularin helps maintain cellular architecture and organelle homeostasis. In skeletal muscle, MTM1 activity is tightly linked to membrane remodeling pathways required for myofiber integrity and excitation–contraction support. Loss-of-function variants in MTM1 are associated with X-linked myotubular myopathy, motivating mechanistic studies of phosphoinositide turnover and muscle cell biology.

myotubularin CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous MTM1 expression without altering the underlying DNA sequence.

myotubularin CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the MTM1 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 MTM1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous myotubularin expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native MTM1 locus and enabling the study of myotubularin-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of myotubularin pathway restoration in tumor cells with silenced or reduced MTM1 expression.

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