WTAP CRISPR Activation Plasmid (m): sc-425635-ACT

Wtap encodes WTAP, a core component of the N6-methyladenosine (m6A) RNA methyltransferase complex that cooperates with METTL3/METTL14 to regulate pre-mRNA processing, alternative splicing, mRNA stability, and translation. WTAP supports nuclear speckle organization and coordinates post-transcriptional gene regulation programs that influence cell-cycle progression, lineage specification, and stress responses. In mouse models, perturbation of WTAP-linked m6A machinery affects embryonic development and tissue homeostasis, reflecting its broad role in gene expression control. Dysregulated m6A pathway activity involving WTAP has been associated with altered proliferative signaling and differentiation states relevant to cancer biology and other diseases driven by aberrant RNA metabolism.

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

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

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