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PRODUCT

SpCas9 Nuclease

SpCas9 protein for precise editing

SpCas9 nuclease is a high-quality, recombinant protein that enables robust and reliable genome editing. This widely used CRISPR nuclease, complexed with a synthetic single guide RNA (sgRNA), can achieve high editing efficiencies. It features two nuclear localization signals (2NLS) to enhance its transport into the nucleus for precise gene editing.

  • Enables high-precision gene editing with a blunt-end cut
  • Features two nuclear localization signals (2NLS) for enhanced nuclear transport
  • Complexed with sgRNA to achieve high editing efficiencies
  • Used in a wide range of applications, including cell and gene therapy

Select a Size

SpCas9 Nuclease (300 pmol)

#R20SPCAS9-Sm
Description Deliverables Specifications gRNA Design Data FAQ Resources
Description

SpCas9 Nuclease for Reliable CRISPR Gene Editing

Synthego’s SpCas9 nuclease is engineered for consistent, high-efficiency editing across diverse cell types. It includes two nuclear localization signals (2NLS) to enhance nuclear delivery and is optimized for use with synthetic sgRNA. When paired with Synthego’s Research sgRNA, SpCas9 generates precise blunt-end double-strand breaks approximately three nucleotides upstream of the 5′-NGG-3′ PAM sequence, enabling reproducible editing outcomes and streamlined optimization across applications.

Image
RNP complex schematic of SpCas9. Depicted in the schematic is genomic DNA (light blue), PAM sequence (pink), and predicted cut site locations on genomic DNA (gray scissors). Nuclease is depicted in light green/blue gradient color. The guide RNA or gRNA is comprised of the target sequence (green) and scaffold (dark blue).

Key Features

  • High Efficiency & Precision: Engineered for consistent editing across diverse cell types with strong on-target activity.
  • Enhanced Nuclear Delivery: Two NLS sequences improve transport of SpCas9 into the nucleus for optimal genome editing.
  • Optimized for Synthetic sgRNA: Achieves precise blunt-end double-strand breaks with Synthego Research sgRNA.
  • Reproducible Results: Streamlines optimization and provides predictable performance across applications.
  • Versatile Applications: Ideal for functional genomics, model generation, and early therapeutic development.

Deliverables

Product Number R20SPCAS9
Intended Use This product is intended for research use only
Shipping Conditions Cold Pack
Source E. coli
Purity ≥ 95.0%
Concentration 3.26 mg/mL
10 mg/mL
*Synthego sources materials from multiple qualified suppliers with buffer concentrations within this range.

Specifications

Specification SpCas9
Size 1368 amino acids
PAM Sequence (N = any nucleotide) 5'-NGG-3’
DNA Cleavage Blunt end cut 3nt upstream of PAM sequence
Endonuclease Domains HNH and RuvC
gRNA Length 97 - 103nt
Target Sequence Length 20 nt
Enzyme Class Type IIa CRISPR-Cas system of Streptococcus pyogenes Cas9
Guide RNA

Designing SpCas9 guide RNA

For designing SpCas9 guide RNA(s), use your favorite guide design platform and select SpCas9 nuclease, which has a 5'-NGG PAM sequence.

After you design your SpCas9 guide RNA, use our easy-to-use order interface to order your guides. We will synthesize them resulting in high-quality synthetic guide RNA you complex with our SpCas9 nuclease to achieve your highest editing efficiency.

The following modifications are included on our SpCas9 sgRNA during synthesis:

2'-O-Methyl at 3 first and last bases, 3' phosphorothioate bonds between first 3 and last 2 bases

Data

SpCas9 Engineered for Reliable Editing Performance

Synthego’s SpCas9 nuclease is engineered to deliver consistent, high editing efficiency, helping streamline CRISPR optimization and reduce experimental variability. Its reliable performance supports efficient target editing and enables smooth progression from early research through downstream functional assays.

Image
HEK293T cells were electroporated with 20pmol and 10pmol of SpCas9, serially diluted from an initial 20µM concentration. Sample sets adhered to a 1:3 SpCas9 to sgRNA ratio, achieving >85% editing efficiency.

SpCas9 Optimized for High-Throughput Screening

Our SpCas9 nuclease supports efficient, scalable genome editing across large, high-throughput screening campaigns. Its reliable performance supports efficient target editing and enables smooth progression from early research through downstream functional assays.

Image
A Highthroughput (HTP) screen of the SpCas9 complex with our high-quality synthetic sgRNA electroporated into HEK293T cells targeting 40 genes. Library preparation utilized the Paragon CleanPlex Panel, and subsequent next-generation sequencing (NGS) analysis revealed an editing efficiency of >68%.
FAQ

Answers to Commonly Asked Research SpCas9 Questions

If you have additional questions please connect with a member of our team.

What are the key differences between Research-Grade and GMP SpCas9?
Research-grade Cas9 is optimized for discovery workflows, while GMP SpCas9 meets extensive release criteria (sterility, endotoxin levels, traceability, regulatory documentation) required for translational and clinical programs.

Does this nuclease contain any chemical tags or fusion domains?
No. This is a recombinant SpCas9 with dual NLS sequences for nuclear import. No affinity tags or large fusions that could impair activity.

What PAM requirements and target-site constraints apply when using SpCas9?
When designing guides for SpCas9, consider both target-site sequence and PAM requirements: SpCas9 recognizes a 5’-NGG-3’ PAM immediately downstream of the target. Optimal guides generally use a 20-nt spacer, though 19–21 nt spacers can be appropriate depending on off-target predictions. Additional considerations for selecting the best gRNA include predicted on-target efficiency, off-target risk, and compatibility with your delivery method. Using validated design tools can help identify gRNAs with the strongest predicted performance for your specific application.

Does SpCas9 produce blunt or staggered cuts, and how does this influence repair outcomes?
SpCas9 generates a blunt-ended double-strand break 3 bp upstream of the PAM. This favors NHEJ-driven indel formation but can be combined with HDR donors for precise knock-ins.

What delivery methods are compatible with SpCas9?
SpCas9 can be used in:

Electroporation/nucleofection (most common)

Microinjection

Lipid-based delivery systems

Learn more about CRISPR delivery methods.

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