Sucrase-Isomaltase CRISPR Activation Plasmid (h): sc-401348-ACT

Human SI encodes sucrase-isomaltase, a brush-border membrane glycohydrolase essential for terminal digestion of dietary carbohydrates in the small intestine. The enzyme complex catalyzes hydrolysis of sucrose and α-1,4/α-1,6 linkages in starch-derived oligosaccharides, supporting monosaccharide availability for absorption and epithelial energy metabolism. SI expression and trafficking are tightly linked to enterocyte differentiation programs and apical membrane protein processing within the secretory pathway. Altered SI function or expression is associated with disaccharidase deficiency phenotypes and contributes to variability in intestinal carbohydrate handling that can influence gastrointestinal disease states.

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

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

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