HLA-G CRISPR Activation Plasmid (h): sc-401226-ACT

HLA-G encodes a non-classical MHC class I molecule that modulates immune recognition by engaging inhibitory receptors such as ILT2/LILRB1, ILT4/LILRB2, and KIR2DL4 on NK cells, T cells, and myeloid populations. Its expression supports immune tolerance at the maternal–fetal interface and contributes to regulation of antigen presentation and cytotoxic effector function within the broader MHC class I/NK checkpoint network. HLA-G is frequently studied in contexts of immune evasion and immunosuppressive microenvironments, including cancer biology and chronic inflammatory settings. Altered HLA-G levels and isoform usage have been associated with transplant tolerance, autoimmunity, and pathogen-associated immune modulation, making it a useful target for mechanistic immunology.

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

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

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