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Custom Construction Service of VSV Expression Plasmid Vector Vesicular Stomatitis Virus (VSV) belongs to the family Rhabdoviridae and is a negative-sense single-stranded RNA virus. VSV primarily infects livestock such as cattle, horses, and pigs, causing vesicular lesions in the mouth, hooves, and other areas. Although VSV has minimal impact on humans, it holds significant value as a research tool in virology, immunology, and gene therapy. RGBiotech offers custom construction service for engineering VSV expression plasmid vectors. By choosing our Vesicular Stomatitis Virus (VSV) expression vector construction service, you will obtain a professional, efficient, and high-quality one-stop solution to help promote your scientific research and project progress. If you have any needs, please feel free to contact us at any time. For more information or to inquire about our services, please contact us via admin@rgbiotech.com. We look forward to collaborating with you to advance scientific research and technological innovation. Why Choose RGBiotech? 1) Competitive Gene Synthesis Platform: We possess advanced gene synthesis technology and equipment, which can quickly and accurately synthesize high-quality foreign gene fragments to meet customers' needs for different gene sequences. Whether it is a simple gene fragment or a complex gene combination, we can provide high-quality synthesis services at a competitive price.2) Extensive Vector Engineering Experience: Our team members have many years of experience in vector construction and the operation of Vesicular Stomatitis Virus (VSV). In past projects, we have successfully constructed various types of VSV expression vectors, accumulating rich practical experience. We can deal with various complex technical challenges and provide professional technical support and solutions for customers. 3) Numerous Successful Cases: Relying on a professional technical team and rich experience, we have successfully completed numerous VSV expression vector construction projects, covering multiple fields such as vaccine research and development, cancer treatment, and gene function research. These successful cases not only prove our technical strength but also provide valuable experience for us to further optimize service processes and improve service quality. 4) Strict Quality Control: We use advanced sequencing technology to comprehensively sequence and verify the constructed VSV expression vectors to ensure the accuracy and integrity of the vector sequence. 5) High-Quality Plasmid Products: We can provide endotoxin-free plasmids suitable for transfection of packaging cells, ensuring the efficiency and stability of plasmids in the cell transfection process. Endotoxin-free plasmids can reduce the toxic effects on cells and improve transfection efficiency, providing a strong guarantee for the successful packaging of recombinant VSV. 6) Detailed Reports and Data Support: After the project is completed, we will provide customers with detailed experimental reports, including COA and sequencing verification data, virus packaging results. Customers can obtain strong data support for subsequent research work. Read More Vesicular Stomatitis Virus (VSV) belongs to the genus Vesiculovirus of the family Rhabdoviridae. It is an enveloped, single-stranded negative-sense RNA virus. VSV holds a significant position in the field of virology research and is widely applied in various aspects such as the study of virus pathogenesis, immune responses, and gene therapy vectors.1. Genome Structure of VSV The VSV genome is approximately 11 kb in size and consists of 5 major genes arranged in the order of 3'-N-P-M-G-L-5' which are critical for viral replication, assembly, and infection. The N gene encodes the nucleoprotein, which is responsible for encapsidating the viral genomic RNA to form the ribonucleoprotein complex (RNP). The P gene encodes the phosphoprotein, which interacts with the N protein and the L protein and participates in viral RNA synthesis. The M gene encodes the matrix protein, which plays a crucial role in virus assembly and budding. The G gene encodes the glycoprotein, which is the main antigen on the virus surface and mediates the binding of the virus to host cell receptors and the membrane fusion process. The L gene encodes the RNA-dependent RNA polymerase, which is responsible for the replication and transcription of the viral genome.2. Diseases Caused by VSV VSV mainly infects livestock such as cattle, horses, and pigs, causing vesicular stomatitis. Affected animals develop vesicles in the oral cavity, tongue, lips, muzzle, hooves, and other parts. After the vesicles rupture, ulcers are formed, leading to difficulties in eating, chewing, and swallowing, weight loss, and a decrease in milk production, causing certain economic losses to the livestock industry. VSV occasionally infects humans, the symptoms are usually mild, presenting as flu-like symptoms such as fever, headache, and muscle pain, and generally can self-heal within a few days. Although VSV has low pathogenicity in humans, strict biosafety protocols must be followed during laboratory operations.3. Applications of Recombinant VSV Recombinant VSV has broad applications in various fields, including:1) Basic Research: To study viral replication mechanisms, host immune responses etc. 2) Gene Delivery Tool: VSV can efficiently infect a variety of cell types and introduce foreign genes into host cells, providing a powerful tool for gene function research and gene therapy. 3) Vaccine Development: Recombinant VSV can be used as a vaccine vector. By inserting foreign antigen genes into the VSV genome, it can express and induce an immune response in the body. For example, the Ebola virus vaccine constructed with VSV as a vector has entered the clinical trial stage and achieved good immune effects. 4) Cancer Treatment: Taking advantage of the natural tropism of VSV for tumor cells, recombinant VSV carrying therapeutic genes can be constructed to achieve targeted killing of tumor cells. At the same time, VSV infection of tumor cells can activate the body's anti-tumor immune response and enhance the treatment effect. 4. Features of Recombinant VSV 1) High Infectivity: The G protein of VSV can recognize a variety of cell surface receptors, enabling it to efficiently infect a variety of mammalian cells, including some cell lines that are difficult to transfect.2) Size Limitations for Foreign Inserts: The VSV vector has size limitations for foreign inserts. Generally, VSV can accommodate the insertion of foreign gene fragments of about 1 - 2 kb. However, as the size of the foreign gene fragment increases, the packaging efficiency and stability of the virus may be affected. When the foreign gene fragment exceeds a certain length, it may lead to the inability of the recombinant virus to be normally packaged or a significant decrease in virus viability. Therefore, when designing recombinant VSV, it is necessary to comprehensively consider the size, function of the foreign gene, and its impact on virus characteristics to ensure the obtainment of an efficient and stable recombinant virus. 3) High Immunogenicity: As a virus, VSV can stimulate a strong immune response in the body, which helps to enhance the effects of vaccines and cancer treatments. At the same time, its immunogenicity can also be regulated by genetic engineering methods to meet different application requirements. 4) High Safety: VSV has low pathogenicity in humans, making it safe for laboratory use. In addition, VSV is less stable in the environment and is not easy to spread and diffuse. 5. Packaging Process of Recombinant VSV 1) Vector Construction: Clone the foreign gene into the VSV expression vector to construct a recombinant expression plasmid. At the same time, construct auxiliary (helper) vectors. The auxiliary vector usually contains the key genes required for VSV replication and packaging but does not have the complete ability to package the virus.2) Cell Transfection: Co-transfect the recombinant expression plasmid and the auxiliary vector into the packaging cell line. Commonly used packaging cell lines include BHK-21 (baby hamster kidney cells), etc. These cells can provide various proteins and enzymes required for VSV replication and packaging. 3) Virus Rescue: After culturing the transfected packaging cells for a period of time, the replication, transcription, and translation of the viral genome, as well as the assembly of virus particles, begin to occur inside the cells. By collecting the cell culture supernatant, the recombinant VSV virus solution can be obtained. 4) Virus Purification and Concentration: Purify the virus solution by methods such as ultracentrifugation and column chromatography to remove cell debris, culture medium components, and other impurities. Then, concentrate the virus by techniques such as ultrafiltration to increase the virus titer and meet the needs of subsequent experiments. 5) Titer Determination: Measurement of viral titer using plaque assay or TCID50. 6. Components of VSV Expression Vectors 1) Promoter: Located at a specific position on the vector, it drives the initiation of transcription of the foreign gene. Commonly used promoters include the VSV's own promoter and some strong promoters such as the CMV promoter, etc., to ensure the efficient expression of the foreign gene in host cells.2) Multiple Cloning Site: It contains multiple restriction endonuclease cleavage sites, which is convenient for the insertion of foreign genes. By selecting appropriate cleavage sites, the foreign gene can be accurately cloned into the vector. 3) Terminator: Located downstream of the foreign gene, it terminates the transcription process and ensures the integrity of the transcription product. 4) Elements Related to Virus Replication and Packaging: These elements are crucial for the replication and packaging process of VSV in packaging cells, ensuring the correct assembly and release of recombinant viruses. 5) Selection Marker Gene: Such as antibiotic resistance gene and reporter gene, etc., are used to screen cells that have successfully introduced the vector after cell transfection. By adding corresponding antibiotics to the culture medium, only cells carrying the selection marker gene can survive and grow. 7. Introduction to the Helper Vector The helper vector mainly contains the genes necessary for VSV replication and packaging, such as the N, P, M, G, and L genes, etc., but lacks virus packaging signals or other key elements, making it unable to package into complete virus particles by itself. The role of the helper vector is to provide the necessary protein support for the generation of recombinant VSV when co-transfected with the recombinant expression plasmid into packaging cells, assisting in the replication, transcription of the recombinant virus genome, and the assembly of virus particles. Different helper vector systems may vary in gene composition and design, but the core purpose is to improve the rescue efficiency and virus yield of recombinant VSV. |
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