LIN-28B CRISPR Activation Plasmid (h): sc-402851-ACT

Human LIN28B encodes the RNA-binding protein LIN-28B, a heterochronic regulator that suppresses maturation of the let-7 microRNA family and thereby reshapes post-transcriptional gene regulation programs controlling proliferation, metabolism, and lineage commitment. By modulating let-7-dependent networks, LIN-28B influences developmental timing, stem-like cell states, and cellular reprogramming, intersecting with pathways linked to MYC, RAS/MAPK, and mTOR signaling outputs. Dysregulated LIN28B activity has been associated with altered differentiation and aberrant growth control in multiple disease-relevant contexts, including oncogenic transcriptional programs and pediatric developmental disorders. These properties make LIN28B a useful node for studying miRNA biogenesis, RNA metabolism, and gene regulatory circuitry in human cells.

LIN-28B CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous LIN28B expression without altering the underlying DNA sequence.

LIN-28B CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the LIN28B 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 LIN28B transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous LIN-28B expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native LIN28B locus and enabling the study of LIN-28B-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of LIN-28B pathway restoration in tumor cells with silenced or reduced LIN28B expression.

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