SRY CRISPR Activation Plasmid (h): sc-404342-ACT

SRY (sex-determining region Y) encodes a high-mobility group (HMG) box transcription factor that initiates male sex determination by directing Sertoli cell differentiation and testis development during embryogenesis. SRY binds and bends DNA to regulate gene expression programs, prominently activating SOX9 and coordinating downstream gonadal differentiation pathways that shape reproductive tract development. Dysregulated SRY expression or sequence variation is associated with disorders of sex development, including gonadal dysgenesis and sex reversal phenotypes, and is also used as a lineage- and sex-specific marker in diverse experimental settings. Because SRY functions as a master regulator, modulating its activity provides a direct handle to interrogate transcriptional networks controlling cell fate decisions.

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

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

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