High Quality Hygromycin B with the CAS number 31282-04-9 is a powerful antibiotic that is widely used in molecular biology and cell culture applications. It is produced by Streptomyces hygroscopicus. Hygromycin B acts by inhibiting protein synthesis in both prokaryotic and eukaryotic cells, which makes it a valuable tool for a variety of research purposes.
Fórmula química: C20H37N3O13
Peso molecular: 527.52 g/mol
Apariencia: White to off - white powder
Solubilidad: Hygromycin B is soluble in water. It is recommended to prepare stock solutions at a concentration of 50 - 100 mg/mL in sterile water.
Condiciones de almacenamiento: Store at -20°C. Avoid repeated freeze - thaw cycles.
1. Cell Selection in Transfection: Hygromycin B is commonly used as a selection agent in cell culture. When cells are transfected with a plasmid containing the hygromycin resistance gene (hygr), only the cells that have successfully taken up the plasmid and express the resistance gene can survive in the presence of hygromycin B. This allows researchers to isolate and culture the transfected cells.
2. Gene Expression Studies: It can be used to maintain the expression of genes in cells over time. By continuously culturing cells in the presence of hygromycin B, only cells that retain the transfected plasmid with the hygromycin resistance gene will survive, ensuring the stable expression of the gene of interest.
3. Microbial Research: Hygromycin B can also be used in microbiological research to study the sensitivity of different microorganisms to the antibiotic. It can help in understanding the mechanisms of antibiotic resistance and the development of new antimicrobial strategies.
1. Preparation of Stock Solution: Weigh an appropriate amount of Hygromycin B powder and dissolve it in sterile water to make a stock solution of 50 - 100 mg/mL. Filter - sterilize the solution using a 0.22 - μm filter to remove any potential contaminants.
2. Determination of Optimal Concentration: The optimal concentration of Hygromycin B for cell selection depends on the cell type. Perform a kill - curve experiment to determine the lowest concentration of Hygromycin B that can kill all non - transfected cells within 1 - 2 weeks. Typical concentrations range from 100 - 500 μg/mL for mammalian cells.
3. Addition to Culture Medium: Once the optimal concentration is determined, add the appropriate volume of the stock solution to the culture medium. Mix well and replace the regular culture medium with the selective medium containing Hygromycin B 24 - 48 hours after transfection.
Case 1: Stable Cell Line Generation
A research group was interested in generating a stable cell line expressing a specific protein for drug screening. They transfected a mammalian cell line with a plasmid containing the gene of interest and the hygromycin resistance gene. After transfection, they added Hygromycin B to the culture medium at a concentration of 200 μg/mL. Over the course of two weeks, non - transfected cells died, and the remaining cells formed colonies. These colonies were then expanded and analyzed for the expression of the target protein. The stable cell line generated using Hygromycin B selection was used successfully for high - throughput drug screening assays.
Case 2: Gene Knock - down Studies
In another study, researchers wanted to study the function of a particular gene by knocking it down using RNA interference. They transfected cells with a plasmid expressing a short hairpin RNA (shRNA) targeting the gene of interest, along with the hygromycin resistance gene. After adding Hygromycin B to the culture medium at 300 μg/mL, they were able to select the cells that had taken up the plasmid. Further analysis showed a significant reduction in the expression of the target gene, demonstrating the effectiveness of Hygromycin B in facilitating gene knock - down studies.
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