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Hygromycin B, EvoPure^®

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Description

Hygromycin B, EvoPure^®is a highly pure form of Hygromycin B (>99.0% pure). Hygromycin B is a unique aminoglycoside antibiotic derived from Streptomyces hygroscopicus and is routinely used as a selection agent in transfection experiments.

TOKU-E Company is able to perform specific tests beyond standard specifications including endotoxin content, arsenic content, cell line testing, spectral analysis, and more.

This product is considered a dangerous good. Quantities above 1 g may be subject to additional shipping fees. Please contact us for questions.

For more Hygromycin B products, click here.

CAS Number

31282-04-9

Molecular Formula

C₂₀H₃₇N₃O₁₃

Molecular Weight

527.52

Mechanism of Action

Hygromycin B inhibits protein synthesis by strengthening the interaction of tRNA binding in the ribosomal A-site. Hygromycin B also prevents mRNA and tRNA translocation by an unknown mechanism. These are unique mechanisms for an aminoglycoside antibiotic and they differ from the mode of action of neomycin, gentamicin, and G418.

Mechanism of Resistance

Hygromycin B resistance is conferred by the hph gene isolated from Streptomyces hygroscopicus , a 1467 bp fragment which encodes hygromycin B phosphotransferase (HPh). Cell lines successfully transfected with the hph gene produce hygromycin B phosphotransferase and convert hygromycin B to 7”-O-phosphoryl-hygromycin B by phosphorylating the 4-hydroxyl group on the cyclitol ring of hygromycin B. 7”-O-phosphoryl-hygromycin B lacks antibiotic activity and does not interact with prokaryotic or eukaryotic ribosomes.

Storage Conditions

2-8°C

Tariff Code

2941.90.1010

Spectrum

Hygromycin B is effective against eukaryotic and prokaryotic cells.

Applications

Eukaryotic Cell Culture Applications

Hygromycin B is routinely used as a selective agent in mammalian cell culture to isolate hygromycin B resistant cells after transfection. Selectable markers for hygromycin B resistant cells include the hyg or hph resistance genes which express a phosphotransferase that inactivates hygromycin B by phosphorylation. Effective working concentrations range from 100 – 1000 µg/mL.

For additional information regarding relevant cell lines, resistance plasmids, and culture media, please visit our cell culture database.

Microbiology Applications

Hygromycin B can be used as a selection agent to isolate Hygromycin B resistant bacteria and fungi. The following Hygromycin B selection concentrations should serve as a guide only and may vary depending on experimental conditions and cells used:

Bacteria (E. coli) - 50 µg/mL - 100 µg/mL
Fungi - 100 µg/mL - 300 µg/mL
Yeasts - 50 µg/mL - 200 µg/mL

Plant Biology Applications

Hygromycin B is routinely used as a selection agent for Arabidopsis plants that have been transformed with a hygromycin B resistance gene. A rapid method to screen for hygromycin B resistant Arabidopsis in less than four days has been developed. After Arabidopsis seeds have been transformed with a resistance plasmid (pBIG-HYG), they are plated on MS medium with hygromycin B and subjected to a two day stratification at 4°C in the dark. Seeds are then exposed to light for 4-6 hours to stimulate germination and then placed in the dark for another two days. Transformed seeds are selected and identified after a 24 hour period in the light. Resistant transformants are characterized by long hypocotyls. (Harrison et al, 2006).

Specifications

Form

Powder

Appearance

Off-white to tan powder

Source

Streptomyces Hygroscopicus

Water Content (Karl Fischer)

≤15%

Melting Point

160-180°C

Assay

|Plasmid_DNA_Transfection_Protocol.pdf|Selection_of_Stable_Transfected_Cell_Lines_Protocol.pdf

Technical Data

Spectral Data - Hygromycin B, EvoPure^®

HNMR Spectra

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Solvent:	D2O
Instrument:	Mercury 300
Frequency:	300 MHz

FTIR Spectra

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Mass Spectra

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Polarity/Scan Type:	Positive
Solvent:	Water
Solution Concentration:	10 mg/mL

References

Dai S et al (2001) Comparative analysis of transgenic rice plants obtained by Agrobacterium-mediated transformation and particle bombardment. Mol. Breeding. 7: 25–33

Harrison S et al (2006) A rapid and robust method of identifying tansformed Arabidopsis thaliana seedlings following floral dip transformation. Plant Methods 2(19):1-7 PMID 17087829

González A, Jiménez A, Vázquez D, Davies JE, Schindler D. (1978) Studies on the mode of action of hygromycin B, an inhibitor of translocation in eukaryotes. Biochim Biophys Acta. 521(2):459-469 PMID 367435