Sar1B CRISPR Activation Plasmid (h): sc-412122-ACT

Human SAR1B encodes Sar1B, a small COPII GTPase that initiates ER-to-Golgi transport by recruiting Sec23/24 and driving vesicle budding from endoplasmic reticulum exit sites. Through regulated GTP binding and hydrolysis, Sar1B coordinates cargo selection and membrane curvature, supporting secretion, lipoprotein trafficking, and broader proteostasis networks linked to ER stress responses. Disruption of SAR1B-dependent COPII dynamics has been associated with congenital disorders of lipid absorption and chylomicron secretion, making it relevant to studies of intestinal lipid handling and systemic lipid homeostasis. Sar1B is also used as a model node for interrogating secretory pathway regulation, membrane trafficking checkpoints, and organelle communication in human cells.

Sar1B CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous SAR1B expression without altering the underlying DNA sequence.

Sar1B CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the SAR1B 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 SAR1B transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Sar1B expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native SAR1B locus and enabling the study of Sar1B-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Sar1B pathway restoration in tumor cells with silenced or reduced SAR1B expression.

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