6CKine CRISPR Activation Plasmid (m): sc-437226-ACT

Mouse Ccl21b encodes the chemokine 6CKine (CCL21), a CCR7 ligand that directs chemotaxis of naïve T cells, central memory T cells, and mature dendritic cells toward lymphatic endothelium and T cell zones in secondary lymphoid organs. By establishing gradients along lymphatic vessels and within lymph nodes, 6CKine supports immune cell trafficking, antigen surveillance, and organization of lymphoid microenvironments. CCL21–CCR7 signaling intersects with GPCR-mediated pathways that regulate actin remodeling, integrin activation, and directional migration, influencing leukocyte positioning and stromal–immune crosstalk. Dysregulated CCL21/CCR7 axis activity has been associated with altered inflammatory cell recruitment, lymphangiogenesis-linked immune remodeling, and microenvironmental changes observed in immune-mediated and neoplastic contexts.

6CKine CRISPR Activation Plasmid (m) provides a targeted, non-destructive approach to upregulating endogenous Ccl21b expression without altering the underlying DNA sequence.

6CKine CRISPR Activation Plasmid (m) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the Ccl21b 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 Ccl21b transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous 6CKine expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native Ccl21b locus and enabling the study of 6CKine-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of 6CKine pathway restoration in tumor cells with silenced or reduced Ccl21b expression.

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