LRG-47 CRISPR Activation Plasmid (m): sc-421030-ACT

Mouse Irgm1 encodes LRG-47, an interferon-inducible immunity-related GTPase that localizes to intracellular membranes and supports cell-autonomous defense against intracellular pathogens. LRG-47 participates in IFN-γ–driven antimicrobial programs by coordinating vesicular trafficking, phagosome maturation, and autophagy-related processes that influence pathogen containment and clearance. It interfaces with innate immune signaling networks that shape macrophage activation states and inflammatory outputs, linking interferon responses to organelle remodeling. Dysregulated Irgm1 activity has been associated with altered susceptibility to infection and immunopathology, making it relevant for studies of host–pathogen interactions and inflammatory disease mechanisms.

LRG-47 CRISPR Activation Plasmid (m) provides a targeted, non-destructive approach to upregulating endogenous Irgm1 expression without altering the underlying DNA sequence.

LRG-47 CRISPR Activation Plasmid (m) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the Irgm1 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 Irgm1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous LRG-47 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native Irgm1 locus and enabling the study of LRG-47-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of LRG-47 pathway restoration in tumor cells with silenced or reduced Irgm1 expression.

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