Orexin-A Double Nickase Plasmid (h): sc-400357-NIC
- Target species: human
- 20 µg of transfection-ready, purified plasmid DNA; Suitable for up to 20 transfections
- Orexin-A Double Nickase Plasmid (h) consists of a pair of plasmids each encoding a D10A mutated Cas9 nuclease and a target-specific 20 nt guide RNA (gRNA) designed to knockout gene expression with greater specificity than its CRISPR/Cas9 KO counterpart
- Paired gRNA sequences are offset by approximately 20 bp to allow for specific Cas9-mediated double nicking of the genomic DNA, which mimics a DSB
- One plasmid in the pair contains a puromycin-resistance gene for selection; the other plasmid in the pair contains a GFP marker to visually confirm transfection
- Orexin-A Double Nickase Plasmid (h) and Orexin-A Double Nickase Plasmid (h2) encode distinct paired gRNA designs targeting HCRT. One or both designs may be available
- Following transfection, gene knockout efficiency can be assayed by WB, IF or IHC using antibody: Orexin-A Antibody (KK09): sc-80263
Ordering Information
| Product Name | Catalog # | UNIT | Price | Qty | FAVORITES | |
Orexin-A Double Nickase Plasmid (h) | sc-400357-NIC | 20 µg | $410.00 | |||
Orexin-A Double Nickase Plasmid (h2) | sc-400357-NIC-2 | 20 µg | $410.00 |
HCRT encodes prepro-orexin, which is proteolytically processed to yield Orexin-A, a neuropeptide primarily produced by hypothalamic neurons that coordinate arousal, sleep–wake stability, energy homeostasis, and motivated behaviors. Orexin-A signals through the G protein-coupled receptors OX1R and OX2R to engage Ca²⁺ flux, MAPK/ERK, and cAMP/PKA pathways, modulating neurotransmitter release across monoaminergic and cholinergic circuits. This signaling network influences feeding and stress responsiveness via hypothalamic and brainstem integration of metabolic and circadian cues. Altered HCRT/orexin tone has been implicated in sleep–wake disorders, obesity-related phenotypes, and neuropsychiatric features, supporting its use in mechanistic studies of neuronal circuit regulation.
Orexin-A Double Nickase Plasmid (h) consists of a matched pair of plasmids engineered for high-specificity editing of the HCRT locus in human cell lines. Each plasmid expresses a Cas9 D10A nickase and a distinct sgRNA targeting opposite DNA strands within HCRT. When directed to adjacent sites on opposite DNA strands, the two nickases generate offset single-strand nicks that together produce a staggered double-strand break, requiring coordinated on-target activity from both guides. The resulting DNA break is resolved by endogenous cellular repair pathways, most commonly through non-homologous end joining (NHEJ), leading to insertions or deletions that disrupt HCRT function. By requiring dual sgRNA engagement at the target locus, the double nicking approach enhances editing specificity and provides a complementary CRISPR strategy for applications where additional control over targeting precision is desired.
To support efficient identification of edited cells, one plasmid encodes GFP for fluorescent visualization of transfected populations, while the companion plasmid carries a puromycin resistance gene for antibiotic selection. Together, these features support efficient enrichment of co-transfected populations and simplify the validation of HCRT-disrupted clones.
For Research Use Only. Not Intended for Diagnostic or Therapeutic Use.