NELF-B CRISPR Activation Plasmid (h): sc-405667-ACT

Human NELFB encodes NELF-B, a core subunit of the Negative Elongation Factor (NELF) complex that cooperates with DSIF to enforce promoter-proximal pausing of RNA polymerase II and shape early transcription elongation. By regulating pause release and transcriptional kinetics, NELF-B influences stimulus-responsive gene programs, enhancer–promoter communication, and broader chromatin-linked control of gene expression. NELF-dependent pausing is tightly connected to pathways governing cell cycle progression, differentiation, and stress responses, making NELFB function relevant to mechanistic studies of transcriptional dysregulation. Altered regulation of elongation control has been implicated across diverse disease contexts, supporting the use of NELFB perturbation to model how pausing dynamics impact downstream phenotypes.

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

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

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