Cas Protein Identification and Characterization Service    

CRISPR-Cas systems, originally part of prokaryote adaptive immunity, have transitioned into powerful tools for eukaryotic genome editing. Central to these systems are Cas proteins, which are crucial for their function. Effective prediction and identification of Cas proteins aid in understanding CRISPR-Cas system types and expanding genome editing capabilities. The discovery of new Cas proteins, like SpCas9, SaCas9, and Cpf1, has propelled CRISPR technology forward, addressing off-target effects and enhancing editing capabilities. Bioinformatics tools have played a pivotal role in this journey, enabling rapid screening of large datasets to identify proteins with similar functions and aiding in deciphering the CRISPR-Cas toolkit. By analyzing spacer sequences, bioinformatics helps determine essential motifs like the protospacer adjacent motif (PAM), crucial for Cas protein function.

Figure 1. Functional classification of Cas proteins. (Makarova KS, et al., 2015)

Overall, bioinformatics-driven approaches lay the foundation for advanced gene editing systems, promising precision and efficiency in genome editing endeavors. By uncovering novel Cas proteins with enhanced specificity, activity, and compatibility, researchers can address existing limitations, such as off-target effects and sequence-based constraints, thereby advancing the development of more precise and efficient gene editing and nucleic acid detection tools.

Our Cas Protein Identification and Characterization Service Process

Creative Biogene's CRISPR/Cas9 platform^CB offers comprehensive Cas protein identification and characterization services tailored to diverse research needs. Through bioinformatic analysis and experimental assessment, optimal Cas proteins are selected and evaluated for activity and specificity. Key properties like thermal stability and nuclease activity are assessed, ensuring suitability for diverse applications and providing a reliable foundation for biotechnological studies. The specific detailed process is as follows:

Figure 2. The workflow of Cas protein identification and characterization.

1. Bioinformatic Analysis for Cas Protein Selection

• Utilize bioinformatics tools to analyze available Cas protein sequences.
• Evaluation of sequence specificity, efficiency, and off-target effects to identify the most suitable Cas protein for the intended application.
• Consideration of factors such as PAM compatibility and target sequence characteristics during the selection process.

2. dsDNA Cleavage Determination

• Generation of Cas protein complexes and their incubation with dsDNA substrates in both cell-free and cell-lysate environments.
• Measurement of dsDNA cleavage activity using quantitative assays such as gel electrophoresis or fluorescence-based assays.
• Comparison of cleavage efficiency and specificity between different Cas protein candidates under simulated intracellular conditions.

3. Determination of PAM Sequence

• Identification and characterization of PAM sequences recognized by the selected Cas protein using bioinformatic prediction tools.
• Experimental validation of PAM sequences through in vitro cleavage assays with synthetic DNA substrates containing various PAM sequences.
• Optimization of PAM sequences to maximize specificity and efficiency of target DNA recognition.

4. Protein Expression and Purification

• Design and construction of expression vectors encoding the selected Cas protein for expression in E. coli host cells.
• Transformation of expression vectors into E. coli cells followed by induction of protein expression using appropriate inducers.
• Purification of expressed Cas proteins using affinity chromatography or other purification techniques to obtain highly pure protein samples suitable for downstream applications.

5. Assessment of Various Properties

• Evaluate key properties including PAM and guide RNA (gRNA) sequences, target sequence length, protein thermostability, and nuclease activity.
• Perform comprehensive analyses to ensure the suitability of the selected Cas proteins for specific applications.

Advantages of Our Cas Protein Identification and Characterization Service

• Prompt Responsiveness to Customer Needs: Our platform ensures swift and efficient responses to customer inquiries and requirements, prioritizing their satisfaction and needs.
• Tailored Personalized Services: We offer customized solutions tailored to each client's specific needs and preferences, ensuring a personalized experience and optimal outcomes.
• Comprehensive Support Ecosystem: Our platform provides a holistic support system, encompassing various resources and assistance to address all aspects of our clients' projects.
• Extensive Expertise in Gene Editing Technology: With a wealth of experience and knowledge in gene editing technology, our team delivers expert guidance and support for diverse genetic editing endeavors.

With over a decade of experience, Creative Biogene's CRISPR/Cas9 platform^CB facilitates rapid Cas protein prediction and continuous optimization of personalized services through ongoing feedback loops. Contact us today to unlock the full potential of CRISPR/Cas9 technology and propel your research forward!

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