Introduction to sgRNA and Cas9 Complex - Ribonucleoprotein (RNP) Synthesis
The successful application of CRISPR genome editing depends on the accurate assembly of the sgRNA-Cas protein complex which forms the ribonucleoprotein (RNP) complex. BOC Sciences specializes in producing superior quality RNP complexes which enable precise gene editing for numerous biological applications. We provide a synthesis service for sgRNA and Cas protein complexes that delivers unmatched stability and efficiency while maintaining compatibility with different cell types and delivery approaches to meet researchers' genome editing requirements.
How do you submit a request for sgRNA and Cas9 complex - ribonucleoprotein (RNP) synthesis?
To initiate your sgRNA and Cas9 complex - ribonucleoprotein (RNP) synthesis request, please follow these steps:
- Provide the sgRNA sequence and the target gene(s) for genome editing.
- Specify the Cas protein variant required (e.g., wild-type, high-fidelity, or nickase).
- Indicate any modifications needed for enhanced stability or delivery efficiency.
- Define the intended application, such as in vitro research, ex vivo cell engineering, or in vivo gene editing.
- Submit your request through the BOC Sciences online platform for a seamless and efficient process.
What is a Ribonucleoprotein (RNP)?
A ribonucleoprotein (RNP) is a molecular complex composed of RNA molecules bound to proteins, playing essential roles in numerous biological processes. In the context of CRISPR-Cas systems, an RNP consists of a guide RNA (sgRNA) and a Cas nuclease, forming a fully functional editing complex that can be directly delivered into cells. Unlike plasmid-based or viral delivery approaches, RNP complexes provide immediate editing activity, reduce the risk of unwanted genomic integration, and improve safety profiles for gene-editing applications.
What is the Function of sgRNA Cas9 complex?
The sgRNA-Cas9 complex is the core machinery of CRISPR-mediated genome editing. The key functions include:
- Target Recognition: The single-guide RNA (sgRNA) directs the Cas9 nuclease to a specific genomic sequence via complementary base pairing.
- DNA Cleavage: The Cas9 enzyme introduces site-specific double-strand breaks (DSBs), enabling precise gene disruption, insertion, or correction.
- Enhanced Efficiency: RNP delivery eliminates the need for cellular transcription and translation, resulting in rapid and transient genome editing activity.
- Reduced Off-Target Effects: Pre-assembled RNP complexes degrade quickly in cells, minimizing prolonged exposure and reducing potential off-target modifications.
This mechanism makes the sgRNA-Cas9 RNP complex a powerful and versatile tool for precise genetic modifications.
Comprehensive Service Guide to sgRNA and Cas9 Complex - Ribonucleoprotein (RNP) Synthesis
BOC Sciences provides a comprehensive suite of services related to the synthesis, optimization, and customization of sgRNA and Cas protein complexes (RNPs) to support diverse genome-editing applications. Our specialized services ensure high efficiency, precision, and scalability for researchers working in functional genomics, gene therapy, and biotechnological applications.
Custom sgRNA Synthesis and sgRNA Modification Service
Our sgRNA synthesis service covers the design, chemical modification, and large-scale production of high-purity, sequence-specific sgRNAs optimized for CRISPR-Cas9, Cas12, and Cas13 systems.
- High-Fidelity sgRNA Synthesis: We provide sgRNAs with stringent quality control, ensuring accurate sequence integrity and high target specificity.
- Chemically Modified sgRNA: Enhancing sgRNA stability and reducing degradation with modifications such as:
(1) 2′-O-methyl (2′-OMe) modifications to increase RNA stability.
(2) Phosphorothioate (PS) linkages to enhance resistance to exonucleases.
(3) Chimeric RNA-DNA modifications to optimize binding efficiency.
- Dual sgRNA Systems: Supporting Cas9-nickase (Cas9n) strategies with two sgRNAs for precise genome editing with minimized off-target effects.
- Long and Truncated sgRNAs: Offering full-length sgRNAs (~100 nt) for maximum binding affinity or truncated sgRNAs (~67 nt) to reduce off-target effects while maintaining high efficiency.
- Multiplexed sgRNA Synthesis: Providing multiple sgRNAs in a single reaction for simultaneous multi-gene editing or genome-wide CRISPR screening applications.
Cas Protein Selection and Customization
We offer an extensive range of Cas nucleases to meet diverse genome-editing needs. Each Cas protein is produced under stringent purification and activity validation protocols, ensuring high enzymatic efficiency and minimal contaminants.
- SpCas9 (Wild-Type Streptococcus pyogenes Cas9): The most widely used Cas9 variant, recognizing the NGG PAM sequence.
- High-Fidelity (HiFi) Cas9: Engineered for significantly reduced off-target cleavage, maintaining high efficiency in gene editing.
- Cas9 Nickase (Cas9n): Mutant Cas9 variants that introduce single-strand breaks, reducing unwanted insertions and deletions
- Cas12a (Cpf1): Recognizing a T-rich PAM, generating staggered DNA cuts, and operating with a single crRNA (without tracrRNA).
- Cas13 Variants: Specialized for RNA targeting and degradation, applicable in transcriptome engineering and viral RNA interference studies.
- dCas9 (Catalytically Inactive Cas9): Used for CRISPR interference (CRISPRi) and CRISPR activation (CRISPRa), enabling gene silencing or transcriptional activation without inducing double-strand breaks.
Optimized sgRNA and Cas9 Complex Assembly & Validation
To maximize genome-editing efficiency, we provide pre-assembled, functionally validated RNP complexes ready for direct delivery into cells. Our RNP assembly services include.
- Custom sgRNA-Cas9 RNP Formation: Pre-loading of sgRNA onto Cas9 under optimized conditions to ensure high binding affinity and stability.
- Cas12 and Cas13 RNP Assembly: Tailored RNP complexes for targeting DNA or RNA, ensuring efficient cleavage and regulatory functions.
- Multiplexed RNP Formation: Simultaneous assembly of multiple RNPs for complex gene-editing strategies, such as gene knock-in, knockout, and large-fragment deletions.
- Enhanced RNP Stability Testing: Assessing optimal storage conditions for lyophilized and liquid RNP formulations to ensure long-term stability.
- Functional Validation: Every batch of RNP undergoes -
(1) Gel shift assays (EMSA) to confirm sgRNA-Cas binding.
(2) In vitro cleavage assays to verify enzymatic activity.
(3) Cell-based editing assays to ensure precise genome modifications.
Step-by-Step Workflow of sgRNA and Cas9 Complex - Ribonucleoprotein (RNP) Synthesis Service
Our streamlined workflow ensures the highest quality and reproducibility of RNP complexes, tailored to meet specific research and application needs.
01
Consultation and Project Discussion
- Requirement Assessment: Our CRISPR experts engage with clients to understand their specific genome-editing needs, including target sequences, application scope, and delivery preferences.
- Design Optimization: We provide professional guidance on sgRNA sequence design, Cas protein selection, and potential chemical modifications to enhance efficiency and stability.
- Customized Solutions: Tailored synthesis plans are developed based on experimental requirements, ensuring optimal performance for in vitro, ex vivo, or in vivo applications.
02
sgRNA Design & Synthesis
- Bioinformatics-driven design ensures high targeting specificity and minimal off-target effects.
- Custom chemical modifications (e.g., 2′-O-methyl, phosphorothioate) available to enhance stability and efficiency.
- High-purity synthesis with rigorous quality control, including HPLC and mass spectrometry analysis.
03
Cas Protein Expression & Purification
- Recombinant Cas proteins (Cas9, Cas12, Cas13, etc.) are expressed in prokaryotic/eukaryotic systems.
- High-yield purification protocols remove endotoxins and ensure enzymatic integrity.
- Activity validation assays confirm functional efficiency before complex formation.
04
RNP Complex Assembly & Optimization
- Optimized sgRNA-Cas protein complexation for maximal genome-editing activity.
- Binding efficiency validation using electrophoretic mobility shift assays (EMSA).
- Stability testing under physiological conditions to ensure prolonged functional activity.
05
Quality Control & Functional Validation
- Analytical assessment of RNP integrity via spectroscopic and electrophoretic methods.
- Genome-editing assays (e.g., in vitro DNA cleavage, cellular transfection) to confirm target-specific activity.
- Batch-to-batch consistency maintained through stringent QC protocols.
06
Packaging & Delivery
- Custom formulations available for various experimental conditions.
- Lyophilized or liquid RNP formats to ensure long-term stability.
- Expedited global shipping with temperature-controlled logistics to maintain product integrity.
With this meticulously optimized workflow, BOC Sciences delivers high-performance, ready-to-use RNP complexes, ensuring superior genome-editing outcomes for diverse research applications.
With this systematic and quality-driven workflow, BOC Sciences guarantees highly efficient and precisely designed pegRNAs that empower researchers to push the boundaries of genome editing.
Key Benefits of Choosing Our sgRNA and Cas9 Complex - Ribonucleoprotein (RNP) Synthesis Service?
Unmatched Expertise
Years of experience in nucleic acid chemistry and protein engineering.
Custom-Tailored Solutions
Fully customized sgRNA sequences and Cas protein formulations for diverse CRISPR applications.
High Purity & Quality Assurance
Rigorous quality control ensures batch-to-batch consistency.
Optimized for Efficiency
Pre-assembled RNPs eliminate the need for in-house preparation, reducing experimental variability.
Scalable & Flexible Production
Capabilities ranging from small-scale research needs to large-scale biopharmaceutical manufacturing.
Global Customer Support
Dedicated technical support for experimental design and troubleshooting.
Applications of sgRNA and Cas9 Complex - Unlocking Genome Editing Potential
Our sgRNA and Cas Protein Complex - Ribonucleoprotein (RNP) Synthesis Service is widely used across multiple research fields, providing precise and efficient genome editing solutions. The versatility and high efficiency of RNP-mediated editing make it an essential tool in various applications, including functional genomics, gene therapy research, synthetic biology, and biopharmaceutical development.
Gene Knockout Studies
RNP complexes facilitate the efficient disruption of specific genes to investigate their function. By introducing targeted double-strand breaks (DSBs), researchers can induce frameshift mutations through non-homologous end joining (NHEJ), leading to gene inactivation. This approach is widely used in studying disease models, regulatory pathways, and cellular functions.
Gene Knock-In and HDR Applications
For researchers aiming to introduce precise modifications, RNPs can be used in conjunction with homology-directed repair (HDR) templates to insert specific DNA sequences at designated genomic loci. This technique is essential for applications such as disease modeling, functional protein tagging, and correction of genetic mutations.
Base Editing and Prime Editing
Beyond traditional CRISPR-mediated DSBs, RNPs are integral to base editing and prime editing technologies, which enable single-nucleotide conversions without introducing breaks in the DNA. This application is particularly beneficial for correcting point mutations associated with genetic disorders or introducing precise modifications in a controlled manner.
Cell Line and Animal Model Generation
Stable gene modifications in mammalian cell lines and animal models are crucial for biomedical research. Our high-purity RNP complexes support efficient genome editing in various systems, including stem cells, primary cells, and in vivo models such as zebrafish, mice, and non-human primates. This application is critical for studying disease mechanisms, drug screening, and regenerative medicine research.
Ex Vivo Gene Editing for Cell Therapy Research
RNP-based gene editing is widely adopted in ex vivo cell engineering, particularly in immune cell therapies (e.g., CAR-T and CAR-NK development). The transient nature of RNP delivery ensures precise modifications without the risk of genomic integration, making it a safer alternative to plasmid or viral-based systems for therapeutic applications.
FAQs about sgRNA and Cas9 Complex - Ribonucleoprotein (RNP) Synthesis
What are the advantages of using RNPs over plasmid-based CRISPR delivery?
RNPs offer higher editing efficiency, faster action, lower off-target effects, and no risk of genomic integration, making them ideal for sensitive applications, including clinical research.
Can you provide a custom sgRNA-Cas RNP complex for my specific target gene?
Yes, we offer fully customized RNP complexes, including target-specific sgRNA design and optimization. Simply provide your target sequence, and we will handle the synthesis and assembly.
How do you ensure the quality and activity of your RNP complexes?
Each RNP complex undergoes rigorous quality control, including sgRNA integrity analysis, Cas protein purity assessment, and functional validation in gene editing assays. This ensures high specificity and efficiency in your experiments.
Can I use your RNP complexes for genome editing in primary cells or stem cells?
Yes, our RNP complexes are optimized for use in various cell types, including hard-to-transfect primary cells and stem cells. We also offer customized formulations to enhance delivery efficiency.
Do you provide different Cas nucleases besides Cas9?
Yes, we offer a range of CRISPR-associated nucleases, such as Cas12a and high-fidelity Cas9 variants. If you need a specific nuclease, we can accommodate custom requests.
Do you offer support for RNP delivery optimization?
Yes, we provide guidance on electroporation, lipofection, and microinjection techniques to maximize RNP delivery efficiency. Our team can also recommend delivery reagents and protocols tailored to your cell type.
BOC Sciences is committed to delivering cutting-edge RNP synthesis solutions with uncompromising quality and efficiency. Contact us today to accelerate your CRISPR research with our high-performance sgRNA and Cas Protein Complex - Ribonucleoprotein (RNP) Synthesis Service.
