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0Description
Hygromycin B is an aminoglycosidic antibiotic produced by Streptomyces hygroscopicus that kills bacteria, fungi and higher eukaryotic cells by inhibiting protein synthesis. It is used for the selection of pro- and eukaryotic cells, stably transfected with the hygromycin resistance gene, as well as for the maintenance of the hygromycin phenotype of resistant cells.
Cloning of the resistance gene and fusion with eukaryotic promoters has resulted in the construction of vectors that allow selection for resistance to Hygromycin B in both prokaryotic and eukaryotic systems.
Application of Hygromycin B
1. Screen and maintain culture of prokaryotic or eukaryotic cells stably transfected with Hph+ vectors
Escherichia coli (Escherichia coli.)-derived hygromycin resistance gene (hyg or hph), encoding hygromycin B phosphotransferase, which converts hygromycin B into a biologically inactive phosphorylated product, thereby detoxifying effect. In view of this principle, hygromycin B is a very useful selectable marker to screen and maintain cultured prokaryotic or eukaryotic cells successfully transfected with hygromycin resistance gene.
2. Hygromycin B is often used in combination with geneticin G418, bleomycin and blasticidin s to screen cell lines stably transfected with two different vectors
The mode of action of hygromycin B differs from that of G418 (Cat#60220ES), Bleomycin (Cat#60257ES) and Blasticidin S (Cat#60218ES), making it ideal for use with another selective combination of antibiotics.
3. Antiviral agent. Hygromycin B selectively penetrates into cells with enhanced permeability due to viral infection, and has the effect of inhibiting translation
4. Add to animal feed as anthelmintic
5. Hygromycin B is added to the culture medium at a concentration that varies with cell type transfected. Recommended concentration for the selection of resistant cells is 50-1000 µg/mL.
For first-time use of the experimental system, it is recommended to establish a kill curve (a dose-response curve) to determine the optimal screening concentration. In general, Commonly used concentrations for selection of various cell types are 50-500 µg/mL for mammalian cells, 20-200 µg/mL for plant cells, bacteria and 200-1000 µg/mL for fungi.
Instructions for Use
1. Plating: When the cells are cultured to 80-90% confluence, discard the medium, wash the cells twice with 1-2 mL PBS, then add 1 mL Trypsin-EDTA solution, wash the cells thoroughly, add an appropriate amount of trypsin digestion solution, and then place the dish at 37ºC for a few minutes to make most of it turn round under the microscope to terminate the digestion. The plating density is about 30% and culture overnight at 37ºC under 5% CO2.
2. Depending on the cell type, set an appropriate concentration gradient. For mammalian cells, it can be set to 50, 100, 250, 500, 750, 1000 μg/mL. Perform the corresponding dilution operation according to the concentration of the stocking solution.
3. Maintain the concentration of Hygromycin B, change the medium every other day, and observe under the microscope. about 7 days later, the lowest concentration of Hygromycin B that completely kills untransformed cells was chosen as the working concentration for the selection of stably transfected cells.
Chemical Properties
Hygromycin B is an off-white to slightly yellowish brown powder with a special odor. It is a weak base, easily soluble in water, methanol, ethanol, etc., and easily forms salts with many organic and inorganic acids.
Storage
The solution can be stored directly at 2℃-8℃ and can be stored for more than 6 months.
Cautions
1. The cells transfected with the hph gene are resistant to hygromycin, and the transfected cells express the gene stably or temporarily.
2. The hygromycin B resistance gene (hyg or hph) was found in other strains besides E.Coli, including Streptomyces hygroscopicus and Klebsiella pneumoniae.
Recommended Products
Hygromycin B (Yeasen) is available in two states, powder (1 g/10 g) and solution (50 mg/mL), with catalog numbers 60225ES and 60224ES, respectively.
|
Product Name |
Cat# |
Size |
|
60224ES03 |
1 g (20 mL) |
|
|
60224ES10 |
10×1 g (20 mL) |
|
Product Name |
Cat# |
Size |
|
Hygromycin B (Inquire) |
60225ES03 |
1 g |
|
Hygromycin B (Inquire) |
60225ES10 |
10 g |
References published by customers using the products
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[3] Huang H, Zhong L, Zhou J, et al. Leydig‐like cells derived from reprogrammed human foreskin fibroblasts by CRISPR/dCas9 increase the level of serum testosterone in castrated male rats[J]. Journal of cellular and molecular medicine, 2020, 24(7): 3971-3981.
[4] Huang H, Zou X, Zhong L, et al. CRISPR/dCas9‐mediated activation of multiple endogenous target genes directly converts human foreskin fibroblasts into Leydig‐like cells[J]. Journal of cellular and molecular medicine, 2019, 23(9): 6072-6084.
[5] Cui Z, Zheng H, et al. Identification and Characterization of the Mitochondrial Replication Origin for Stable and Episomal Expression in Yarrowia lipolytica. ACS Synthetic Biology. 2021 Apr 16;10(4):826-835.
[6] Yao G, Chen X, Han Y, et al. Development of versatile and efficient genetic tools for the marine-derived fungus Aspergillus terreus RA2905[J]. Current Genetics, 2022: 1-12.
[7] Wang W Q, Tang W H. Generation of Fusarium graminearum Knockout Mutants by the Split-marker Recombination Approach[J]. Bio-protocol, 2018: e2976-e2976.
[8] An C, Wang M, Yao W. Exhausting hsa_circ_0072088 restrains proliferation, motility and angiogenesis of breast carcinoma cells through regulating miR-1236-3p and RRM2 in a ceRNA pathway[J]. Clinical Breast Cancer, 2022.
