 |
|
|
| Home -> Products & Services -> Custom Plasmid Vector Construction Services -> Custom Construction Service of Lentiviral Expression Plasmid Vector | ||
| Quick Inquiry |
Custom Construction Service of Lentiviral Expression Plasmid Vector Lentiviral vectors are gene delivery tools derived from modified HIV-1, capable of efficiently infecting both dividing and non-dividing cells. They enable stable integration of exogenous genes into the host genome for long-term expression. Optimized for safety, these vectors are widely used in basic research, gene therapy, and cell engineering. RGBiotech offers custom construction service of lentiviral expression plasmid vectors. If you’d like to get more information regarding our custom construction services, please contact us at admin@rgbiotech.com. Our expert team is ready to help! Why Choose RGBiotech? 1. Flexible and Diverse Backbone Vectors: We have a rich library of lentiviral backbone vectors, which can meet different experimental needs. Including but not limited to the pLVX series, pLenti series, etc. 2. Abundant Promoter Options: Provide a variety of promoters, such as CMV, EF1α, CAG, U6 or H1 (for shRNA), inducible as well as tissue specific promoters (e.g., neuronal Synapsin, liver-specific ApoE), etc. The appropriate promoter can be selected according to the experimental purpose and cell type to precisely regulate gene expression. 3. Multiple Reporter Genes: Include reporter genes such as EGFP, mCherry, Luciferase, etc., which are convenient for monitoring virus infection and gene expression. 4. Mammalian Resistance Selection Markers: Such as Puromycin, Neomycin, Hygromycin, etc., can be used to screen cell lines stably expressing foreign genes. 5. Providing Lentiviral Packaging Plasmid Vectors: In addition to the conventional lentiviral expression vector construction service, we also provide lentiviral packaging plasmid vectors, which can be used to package and produce lentiviral particles with different types of envelope proteins. In addition to the common VSVG envelope protein, it also includes envelope proteins such as 10A1, GALV, RD114, etc., which can expand the host range of the virus and meet special experimental needs. 6. Free Design Optimization: Sequence analysis, codon optimization, and element compatibility guidance. 7. Rigorous Quality Control: Sequencing verification, endotoxin removal upon request. 8. Cost-Effective Pricing: Bulk order discounts. Read More 1. Introduction Lentiviral vectors belong to the Retroviridae family. Due to their ability to integrate the carried foreign genes into the host cell genome after infecting both dividing and non - dividing cells, achieving long - term stable expression, they are widely used in fields such as gene therapy and gene function research. Derived from the human immunodeficiency virus (HIV), after a series of modifications, the pathogenic genes have been removed, while the key elements for viral infection and integration have been retained, making them safe and effective gene delivery tools.2. Applications of Lentiviral Vectors 1) Gene Therapy: Used to introduce normal genes or therapeutic genes (e.g., CRISPR/Cas9, shRNA, CAR genes) into patients to correct genetic defects and treat cancers, single - gene genetic diseases such as hemophilia and sickle - cell anemia.2) Cell Therapy: Modify immune cells, such as in CAR - T cell therapy, to enhance the killing ability of immune cells against tumor cells. 3) Gene Function Research: By overexpressing or knocking down specific genes, study the functions of genes in processes such as cell growth, differentiation, and metabolism. 4) Neuroscience Research: Can be used to label and track neurons and study the structure and function of neural circuits. 5) Stable Cell Line Construction: Rapid establishment of mammalian cell lines stably expressing exogenous genes. 6) Organoid and In Vivo Studies: Support gene delivery in vitro and in vivo for disease modeling and drug screening. 3. Features of Lentiviral Vectors 1) Efficient Infection: Capable of efficiently infecting a variety of cell types, including primary cells and cells that are difficult to transfect.2) Stable Integration: Integrate foreign genes into the host cell genome, achieving long - term stable expression and avoiding the loss of gene expression. 3) Low Immunogenicity: After modification, the immunogenicity of the virus is greatly reduced, reducing the rejection reaction of the host immune system to the vector. 4) Can Carry Large Fragments: Able to carry relatively large foreign gene fragments, meeting the needs of various gene manipulation requirements. 4. Introduction to Different Generations of Lentiviral Vectors 1) First - Generation Lentivirus: The earliest developed lentiviral vector system contains the basic elements of HIV, such as gag, pol, env and other genes. However, due to the risk of generating replication - competent viruses (RCR), the safety is low.2) Second - Generation Lentivirus: The first - generation vector was improved. Some HIV genes were removed, and its functional elements were dispersed into multiple plasmids, reducing the risk of RCR generation. At the same time, the safety and packaging efficiency of the vector were improved. 3) Third - Generation Lentivirus: Further optimized, a self - inactivating (SIN) vector design was adopted. Deletions were introduced into the long terminal repeat sequence (LTR) of the vector, causing the vector to lose transcriptional activity after integrating into the host genome, greatly improving safety. In addition, some regulatory elements such as the internal ribosome entry site (IRES) were added to improve the efficiency and controllability of gene expression. 4) Fourth - Generation Lentivirus: Based on the third - generation, the packaging system was further optimized, increasing the virus titer and packaging efficiency. At the same time, a variety of new envelope proteins were developed, expanding the host range and targeting of the virus. 5. Key Elements of Lentiviral Expression Vectors 1) Packaging Signal (Ψ): A cis - acting element necessary for virus packaging. It can be recognized by virus packaging proteins and package vector RNA into virus particles.2) Long Terminal Repeat Sequence (LTR): Located at both ends of the vector, containing regulatory elements such as promoters and enhancers, responsible for the transcription and integration of the vector. 3) RRE/Rev Response Elements: Facilitate nuclear export of unspliced RNA. 4) cPPT/CTS: Central polypurine tract for enhanced nuclear import. 5) Foreign Gene Expression Cassette: Contains a promoter, a foreign gene, and a terminator. The promoter drives the expression of the foreign gene, and the terminator ensures the correct termination of transcription. 6) Selection Marker: Such as antibiotic resistance genes, used to screen cells containing the vector. 6. Packaging Limit of Lentivirus Lentiviral vectors can usually package fragments of 5 - 8kb. However, with the development of technology, through optimizing the vector design and packaging system, in some cases, larger fragments can be packaged, up to about 10kb at most. However, the larger the packaged fragment, the lower the packaging efficiency may be, and it needs to be evaluated and optimized according to specific circumstances.7. Packaging Process of Lentiviral vectors 1) Vector Construction: Clone the foreign gene into the lentiviral expression vector to construct a recombinant vector.2) Cell Culture: Culture the cell line used for virus packaging, such as HEK293T cells, to ensure good cell status. 3) Transfection: Co – transfection of the expression vector and packaging plasmids into packaging cells. Common transfection methods include liposome transfection and calcium phosphate transfection. 4) Virus Harvest: At a certain time after transfection (e.g. 48h), collect the cell culture supernatant containing virus particles. 5) Virus Concentration: Concentrate the virus supernatant by methods such as ultracentrifugation and ultrafiltration to increase the virus titer. 6) Virus Titer Determination: Use methods such as quantitative PCR and fluorescence microscopy observation to determine the virus titer and determine the infectivity of the virus. |
|
|
Products & Services Resources | ||
|
Home Products & Services Support About Us Contact Us |
Promotions Flyers Brochures Publications News & Events |
Terms & Conditions Privacy Disclaimers |
Contact Us EMAIL: admin@rgbiotech.com |
||
| © RGBiotech All Rights Reserved. | |||||