CrkRS CRISPR Activation Plasmid (h): sc-402238-ACT

Human CDK12 encodes a cyclin-dependent kinase that partners with Cyclin K to phosphorylate the RNA polymerase II CTD, coordinating transcription elongation with co-transcriptional RNA processing. CDK12 is particularly important for sustaining expression of long, complex genes involved in DNA damage response, replication stress tolerance, and maintenance of genome stability, linking it to homologous recombination and checkpoint signaling networks. Perturbation of CDK12 function alters transcriptional programs that govern cell-cycle progression and repair pathway choice, with downstream effects on chromosomal integrity and cellular fitness. In biomedical research, CDK12 is frequently studied in the context of dysregulated transcription–repair coupling and its association with genomic instability phenotypes observed across multiple disease models.

CrkRS CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous CDK12 expression without altering the underlying DNA sequence.

CrkRS CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the CDK12 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 CDK12 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous CrkRS expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native CDK12 locus and enabling the study of CrkRS-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of CrkRS pathway restoration in tumor cells with silenced or reduced CDK12 expression.

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