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| Home -> Products & Services -> Custom Plasmid Vector Construction Services -> Custom Construction Service of Non-Viral Mammalian Gene Expression Plasmid Vector | ||
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Custom Construction Service of Non-Viral Mammalian Expression Plasmid Vector Non-viral mammalian gene expression plasmid vectors do not contain any viral elements and are widely used in both research and gene therapy field. RGBiotech offers custom services to help our clients to construct fit-for-project non-viral mammalian expression plasmid vectors. We have many years of experience in the field of vector construction, and possesses a professional technical team and advanced experimental equipment. The members of our technical team all have a profound background in molecular biology, and we have accumulated a large number of successful cases in plasmid vector construction. If you have the need for mammalian non - viral expression plasmid vector construction, please feel free to contact us at admin@rgbiotech.com. Why Choose RGBiotech? 1. Personalized Customization: According to the different needs of customers, personalized construction schemes for mammalian non - viral expression plasmid vectors are provided to ensure that the special requirements of each experiment are met. 2. High - efficiency and Fast Service: The optimized experimental process and advanced technical platform can complete the vector construction task quickly while ensuring quality, saving precious time for customers. 3. Strict Quality Control: From experimental design, raw material procurement to experimental operation and result verification, every link strictly follows the standard operating procedures (SOP) to ensure the reliable quality of the constructed plasmid vectors. The constructed plasmid vectors are fully sequenced and compared with the designed sequences to ensure the accuracy of the base sequence and avoid problems such as base mutations or deletions. Read More In the field of life science research, gene delivery technology has always been the key to research. Mammalian non - viral expression plasmid vectors, as important gene delivery tools, have received increasing attention in recent years. Compared with viral vectors, they have unique advantages and show broad application prospects in many aspects such as gene therapy, drug research and development, and cell biology research. 1. Applications 1) Gene TherapyBy introducing therapeutic genes into patient cells to repair or replace defective genes, it brings new treatment hopes for many genetic diseases and difficult diseases. For example, in the treatment research of some single - gene genetic diseases, using non - viral expression plasmid vectors to introduce normal genes into patient cells is expected to achieve a radical cure of the disease. 2) Drug Research and Development In the process of drug research and development, in - depth research on drug targets is required. By constructing non - viral expression plasmid vectors and introducing related genes into cells, the effects of drugs on cell physiological functions and gene expression can be studied, accelerating the drug research and development process. 3) Cell Biology Research It helps researchers to deeply understand the physiological processes and signal transduction pathways of cells. For instance, when studying the role of a specific gene in cell cycle regulation, non - viral expression plasmid vectors can be used to introduce the gene into cells for observation and analysis. 2. Delivery Pathways of Exogenous Genetic Material There are mainly the following pathways to deliver exogenous genetic material into cells through non - viral expression vectors.1) Chemical Transfection Method Using chemical reagents such as cationic liposomes and cationic polymers to form complexes with plasmid DNA, which enter cells through the endocytosis of the cell membrane. This method is relatively simple to operate and has a low cost, but it has certain toxicity to cells, and the transfection efficiency is affected by many factors. 2) Physical Transfection Method Including electroporation, microinjection, etc. Electroporation forms small pores on the cell membrane through the action of an electric field, allowing plasmid DNA to enter cells; microinjection directly injects exogenous genetic material into cells. The transfection efficiency of the physical transfection method is relatively high, but it has high requirements for equipment and operation techniques, and may cause greater damage to cells. 3. Common Elements 1) Promotersa) CMV Promoter: It has strong transcriptional activity and can efficiently initiate gene expression in a variety of mammalian cells, and is widely used in various gene expression studies. b) EF1α Promoter: It is stably expressed continuously in most mammalian cells and is often used in experiments that require long - term stable expression of exogenous genes. 2) Reporter Genes a) GFP (Green Fluorescent Protein): It emits green fluorescence and is easy to observe through a fluorescence microscope, which can intuitively detect gene expression and cell transfection efficiency. b) Luciferase: By catalyzing the luminescence of luciferin substrates, it can be quantitatively detected with high sensitivity and is often used in research on gene expression regulation and drug screening. 3) Bacterial Resistance Selection Markers a) Ampicillin Resistance Gene: It enables cells containing the plasmid to grow in a medium containing ampicillin, which is convenient for screening positive clones. b) Kanamycin Resistance Gene: It is also used to screen cells containing the corresponding plasmid and is widely used in genetic engineering experiments. 4) Mammalian Resistance Selection Markers a) Puromycin: Puromycin is a powerful antibiotic - based selective agent in mammalian cell culture. It exerts its effect by binding to the ribosomal A - site and prematurely terminating translation during protein synthesis. When cells are transfected with a plasmid containing the puromycin resistance gene (pac), they can produce an enzyme that inactivates puromycin. This allows the transfected cells to survive in a culture medium supplemented with puromycin, while non - transfected cells are killed. It is widely used in gene expression studies, gene knockout experiments, and stable cell line generation due to its relatively fast - acting nature and high selectivity. b) Neomycin: Neomycin is another commonly used antibiotic for mammalian cell selection. The neomycin resistance gene (neo) encodes an enzyme called neomycin phosphotransferase. This enzyme phosphorylates neomycin, rendering it inactive. In cell culture, when cells are transfected with a vector carrying the neo gene, they gain resistance to neomycin - related antibiotics such as G418. Neomycin - based selection is often employed in a variety of applications, including the establishment of transgenic cell lines, as well as in gene transfer and expression studies. It provides a reliable way to isolate and maintain cells that have successfully incorporated the desired genetic material. c) Hygromycin: Hygromycin is an aminoglycoside antibiotic used for selecting transfected mammalian cells. The hygromycin resistance gene (hph) codes for an enzyme that modifies hygromycin, thereby preventing its inhibitory effect on protein synthesis. Cells transfected with a plasmid containing the hph gene can survive in a culture medium containing hygromycin, while non - transfected cells are eliminated. Hygromycin selection is particularly useful in situations where other selection markers may not be suitable, and it is often used in combination with other techniques to create stable cell lines with specific genetic modifications. d) Blasticidin: Blasticidin is a nucleoside - antibiotic used for the selection of transfected mammalian cells. The blasticidin resistance gene (bsd) encodes a deaminase enzyme that inactivates blasticidin. When cells are transfected with a vector carrying the bsd gene, they can resist the cytotoxic effects of blasticidin in the culture medium. Blasticidin - based selection is relatively quick and efficient, and it is often utilized in experiments involving the generation of stable cell lines, especially when dealing with cells that are difficult to select using other antibiotics. It offers an alternative option for researchers to isolate and propagate cells with the desired genetic constructs. 4. Advantages and Disadvantages of Non - viral Expression Vectors 1) Advantagesa) High Safety: It does not contain viral pathogenic genes, avoiding the risks of immune responses and insertional mutations that may be caused by viral vectors, and greatly improving the safety of gene therapy and research. b) Large - fragment Sequence Insertion Tolerance: It can carry large exogenous gene fragments, meeting the needs of some complex gene structure and function studies. For example, in some gene therapy programs, longer therapeutic gene sequences need to be introduced, and non - viral expression plasmid vectors can well meet this requirement. 2) Disadvantages The delivery efficiency is relatively low. Compared with viral vectors, the efficiency of non - viral expression vectors entering cells is relatively low, which to some extent limits their application scope. However, with the continuous development of technology, new delivery methods and materials are constantly emerging, and it is expected to gradually improve their delivery efficiency. |
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