ALY CRISPR Activation Plasmid (h): sc-403711-ACT

Human ALYREF encodes ALY, an RNA-binding adaptor of the TREX complex that couples co-transcriptional mRNA processing to nuclear export. ALY coordinates recruitment of the NXF1/TAP export receptor and participates in spliceosome-associated steps that ensure efficient maturation and trafficking of polyadenylated transcripts, linking transcription, splicing, and export. Through its roles in RNA metabolism and genome-wide gene expression control, ALYREF is frequently studied in pathways governing cell proliferation, differentiation, and stress responses. Dysregulation of TREX-dependent mRNA export and ALYREF expression has been reported in multiple cancer contexts and other disorders with altered RNA processing, motivating mechanistic studies of how export factors shape disease-associated transcriptomes.

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

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

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