Services
Providing Advanced Biological Technical Services for Researchers in Fundamental Science
CrY2H-seq Library Screening Service
CrY2H-seq Library Screening: High-Throughput Protein Interaction Screening Using Cre-LoxP Technology
CrY2H-seq (Cre-LoxP Yeast Two-Hybrid Sequencing) is an advanced, high-throughput protein interaction screening technology based on the yeast two-hybrid (Y2H) library platform. Designed to enable large-scale protein-protein interaction discovery, CrY2H-seq utilizes the precision of Cre recombinase to target and recombine LoxP sequences, offering effective and high-accuracy interaction screening.
What is CrY2H-seq?
CrY2H-seq is a powerful screening method that leverages the Cre-LoxP system to detect protein-protein interactions within a yeast two-hybrid system. The Cre recombinase enzyme specifically recognizes LoxP sequences, catalyzing the recombination of DNA to identify interactions between proteins, enabling high-throughput, precise screening of protein interaction networks.
How CrY2H-seq Works
The fundamental steps in large-scale protein interaction screening using the Cre-LoxP system are as follows:
Construction of the Cre-LoxP Vector: A bidirectional fusion protein expression vector containing two library sequences to be screened and LoxP sequences is constructed.
Protein Interaction Screening: The Cre-LoxP vector is introduced into a yeast two-hybrid system. When two proteins interact, Cre recombinase will catalyze the recombination of LoxP sequences, generating a new DNA fragment.
Selection and Identification: Protein interactions are screened using yeast two-hybrid, followed by high-throughput sequencing and data analysis to identify interacting proteins.
This process allows the efficient identification of protein-protein interactions, providing valuable insights into protein functions and cellular networks.
Technical Advantages of CrY2H-seq
High-Throughput Screening: The Cre-LoxP system allows simultaneous detection of multiple protein interactions, significantly increasing screening efficiency.
High Accuracy: With its high specificity, the Cre-LoxP system provides more reliable and precise protein interaction data.
Broad Applicability: Compatible with various cell types and biological systems, enhancing its versatility across different research areas.
Integration with Third-Generation Sequencing: Combined with self-activation verification, CrY2H-seq improves the positive interaction rate, ensuring more accurate results.
Service Overview
At Nanjing ProNet Biotech, we specialize in protein interaction validation and have developed a robust yeast two-hybrid experimental system. Building on this foundation, we now offer the CrY2H-seq service—an innovative protein interaction validation method based on the Cre-LoxP principle. This service enables high-throughput protein-protein interaction screening within a yeast two-hybrid library framework.
The Cre-LoxP system is widely recognized in gene editing for transgenic animal models. By utilizing this technology for protein interaction screening, we ensure efficient and accurate identification and validation of protein interactions.
Sample Requirements
To ensure the best results, please provide samples according to the following guidelines:
Sample Type | Sample Requirements | Storage Conditions | Shipment Conditions |
Cell Samples | Cell Count>1*108 | -80°C | Dry ice |
Animal Samples | >5g | -80°C | Dry ice |
Plant Samples | >5g | -80°C | Dry ice |
Total RNA | >400 ug | -80°C | Dry ice |
CrY2H-seq Library-to-Library Interaction Screening Service Workflow & Deliverables
CrY2H-seq Librar.y-to-Library Interaction Screening | |||
Items | Service Content | Working day | Deliverables |
Yeast Library Construction | 1. RNA extraction 2. mRNA extraction | 5 | 1. Primary and Secondary Library Glycerol Stocks 4 ml (Divided into 4 Tubes) 2. Primary Library (2 tubes > 200 µg) 3. Word Document Electronic Version of the Project Report (Including Experimental Data) 4. The Library is Committed to Meeting the Following Criteria: Library Capacity > 1*107CFU Average Insert Fragment Size 0.8-1.5 kb (varies by species) 5. Empty Vector Rate < 5% |
3. cDNA first strand synthesis 4. cDNA second strand synthesis 5. cDNA adapter preparation 6. Recombination of cDNA with attB1 linker. 7. cDNA Stratification and Separation 8. BP Recombination (pDONR222) 9. Electroporation of E. coli DH10B 10. Library Quality Control and Plasmid Extraction 11. Library Plasmid Recombination (pDBlox, pADlox) 12. Electroporation of E. coli DH10B 13. Library Plasmid and Plasmid Extraction | 20 | ||
Exclusion of self activation in the Bait Library | 14. Library Competent Cell Preparation: pADlox Library Plasmid, pDBlox Library Plasmid, and pADlox Empty Vector Transformation into Yeast 15. Yeast Colony PCR Identification 16. Bait Library Mating 17. Bait Library Third-Generation Sequencing | 30 | 1. Raw Data of Bait Library Third-Generation Sequencing 2. Word Document Electronic Version of the Project Report (Including Experimental Data) |
Yeast CrY2H Library Screening | 18. Yeast CrY2H Library Screening 19. First-Generation Sequencing of 96 Positive Clones 20. Third-Generation Sequencing 21. Data Analysis and Organization, Interaction Network Diagram, Final Report | 30 | 1. At least 15 positive clone sequences 2. Raw Data of Library Screening Third-Generation Sequencing 3. Interaction Network Diagrams 4. Word Document Electronic Version of the Project Report (Including Experimental Data) |
Case Study
Positive and Negative Validation: Using the Cre-LoxP screening system, we successfully validated protein interactions through the identification of the Lox77 recombination site with a 5' large T sequence and a 3' p53 sequence.
Library-to-Library Protein Interaction Screening: By applying CrY2H-seq to multiple protein libraries, we conducted large-scale interaction screening and generated sequencing data for identifying interacting proteins and constructing interaction networks.
Why Choose Us?
Expertise: Over years of experience in protein interaction screening and validation.
Cutting-Edge Technology: We use state-of-the-art Cre-LoxP-based technology to ensure high-quality and reliable results.
Tailored Services: Our services are highly customizable to meet the specific needs of your research.
Global Reach: We provide services to researchers and institutions worldwide.
Contact Us
Whether you're exploring novel protein interactions or seeking high-throughput screening solutions, our CrY2H-seq service provides the precision and scalability you need to drive your research forward.
Get in Touch Today to discuss your project and learn how we can help you achieve reliable, high-quality results with our advanced Cre-LoxP-based technology.
Contact us now to schedule a consultation or request a quote!
Disclaimer: All products and services provided are for research purposes only and not for diagnostic or therapeutic use.
Services Workflow
Online Consultation
01
Solution Matching
02
Service Contract
03
One-Stop-Services
04
Project Report
05
Related Products
Product Inquiry
Contact Us
Tel: +86 025-85205672
Email: info@pronetbio.com
Address: Building 3C, Nanjing Xianlin Zhigu,
Qixia District, Nanjing, Jiangsu, China, 210033
Need more information?
Let's connect!