DAT/Dopamine Transporter/SLC6A3 CRISPR Activation Plasmid (h): sc-400391-ACT

SLC6A3 encodes the dopamine transporter (DAT), a presynaptic plasma membrane symporter that clears dopamine from the synaptic cleft by Na⁺/Cl⁻-dependent reuptake, thereby shaping dopaminergic tone and signal duration. DAT regulates vesicular recycling and dopamine homeostasis in nigrostriatal and mesocorticolimbic circuits, influencing reward processing, locomotion, and executive function. Altered SLC6A3 expression or transporter activity has been associated with dysregulated dopaminergic neurotransmission implicated in neuropsychiatric and neurodegenerative research contexts, including attention-related phenotypes, substance use biology, and Parkinsonian pathway vulnerability. DAT also interfaces with kinase-dependent trafficking and phosphorylation programs that control membrane localization and uptake capacity.

DAT/Dopamine Transporter/SLC6A3 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous SLC6A3 expression without altering the underlying DNA sequence.

DAT/Dopamine Transporter/SLC6A3 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the SLC6A3 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 SLC6A3 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous DAT/Dopamine Transporter/SLC6A3 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native SLC6A3 locus and enabling the study of DAT/Dopamine Transporter/SLC6A3-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of DAT/Dopamine Transporter/SLC6A3 pathway restoration in tumor cells with silenced or reduced SLC6A3 expression.

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