Blasticidin

Selection antibiotic: endotoxin tested, sterile reagent

Blasticidin is an efficient selection antibiotic that acts on both eukaryotic and prokaryotic cells. 

Blasticidin is a peptidyl nucleoside antibiotic isolated from Streptomyces griseochromogenes that inhibits protein synthesis by interfering with the peptide-bond formation in the ribosomal machinery.

Resistance to blasticidin is conferred by the blasticidin resistance gene from Bacillus cereus (bsr), which codes for blasticidin-S deaminase [1].
Resistance to blasticidin is also conferred by the blastcidin S acetyltransferase gene (bls) from Streptoverticillum sp [2], and the blasticidin S deaminase gene (BSD) from Apergillus terreus [3].

Typically, mammalian cells are sensitive to blasticidin concentrations of 1-10 µg/ml, and bacteria to 25-100 µg/ml.
Cell death induced by blasticidin occurs rapidly, allowing for selection of transfected cell lines carrying a blasticidin resistance gene within one week. 

References:

1. Izumi M. et al., 1991. Blasticidin S-resistance gene (bsr): A novel selectable marker for mammalian cells. Exp.Cell Res.197:229-33.
2. Perez-Gonalez J. et al., 1990. Cloning and characterization of the gene encoding a blasticidin S acetyltransferase from Streptoverticillum sp. Gene. 86:129-34. 
3. Kimura M. et al., 1994. Blasticidin S deaminase gene from Aspergillus terreus (BSD): a new drug resistance gene for transfection of mammalian cells. Biochim. Biophys. Acta. 1219:653-9.

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Blasticidin by InvivoGen

Specifications

Product concentration: 10 mg/ml

Quality Control: Each lot is thoroughly tested to ensure the absence of lot-to-lot variation.

Purity: ≥ 95% (HPLC)

Endotoxin level: < 1 EU/mg

Physicochemical characterization: pH, appearance

Cell-culture tested: potency validated in blasticidin-sensitive and blasticidin-resistant mammalian cell lines

Non-cytotoxicity of trace contaminants: absence of long-term effects confirmed in blasticidin-resistant cells

Contents

Blasticidin hydrochloride is supplied as a sterile filtered solution at 10 mg/ml in HEPES buffer.

This product is available in four pack sizes:

• ant-bl-05: 5 x 1 ml (50 mg)
• ant-bl-1: 10 x 1 ml (100 mg)
• ant-bl-5: 50 x 1 ml (500 mg)
• ant-bl-5b: 1 x 50ml (500 mg)

Blasticidin hydrochloride is also supplied as a powder:

• ant-bl-10p: 1 g (powder)

Blasticidin is shipped at room temperature.
Upon receipt it should be stored at 4°C or -20°C.
Blasticidin is a harmful compound. Refer to safety data sheet for handling instructions.

Details

WORKING CONCENTRATIONS

The working concentration of blasticidin for mammalian cell lines varies from 1 to 10 μg/ml, in a few cases up to 30 μg/ml.
In a starting experiment, we recommend determining optimal concentrations of antibiotic required to kill your host cell line.
After treatment, cell death occurs rapidly, allowing the selection of transfected cells with plasmids carrying the bsr or BSD genes in as little as 7 days post-transfection.
Suggested concentrations of blasticidin for selection in some examples of mammalian cells are listed below.

References:

1. Dorgham K. et al., 2009. An engineered CX3CR1antagonist endowed with anti-inflammatory activity. J Leukoc Biol. 86(4):903-11.
2. Le Bon L. et al., 2014. Fringe proteins modulate Notch-ligand cis and trans interactions to specifysignaling states. eLife Sci, 3:e02950.
3. Tomecki R. et al., 2014. Multiple myeloma-associatedhDIS3 mutations cause perturbations in cellular RNA metabolism and suggest hDIS3 PIN domain as a potential drug target. Nucleic Acids Res. 42:1270-90.
4. Edbauer D. et al., 2004. Co-expression of nicastrin and presenilin rescues a loss of function mutant of APH-1.J Biol Chem. 279:37311-5.
5. Khandelia P. et al., 2011. Streamlined platform for short hairpin RNAinterference and transgenesis in cultured mammalian cells. PNAS 108:12799-804.
6. Lee HK.et al., 2007. Application of beta-lactamase enzyme complementation to the high-throughputscreening of toll-like receptor signaling inhibitors. Mol Pharmacol. 72:868-75.
7. Matsumoto G. et al., 2011. Serine 403 phosphorylation of p62/SQSTM1 regulates selective autophagicclearance of ubiquitinated proteins. Mol Cell. 44:279-89.
8. Schepetkin IA. et al., 2009.Immunomodulatory activity of oenothein B isolated from Epilobium angustifolium. J Immunol. 183:6754-66.

FAQs

What is the preferred storage method for Blasticidin?
Blasticidin is shipped at room temperature. Upon receipt, it should be stored at 4°C for short term storage or -20°C for long term storage. Avoid repeated freeze-thaw cycles. The expiry date is specified on the product label.
Once Blasticidin solution opened, it remains stable for 1 year at -20°C providing you avoid repeated freeze-thaw cycles, and it can be kept at 4°C for 3 months provided it is kept in sterile conditions.
Specifically, for Blasticidin powder, the stock solution must be at a concentration of 10 mg/ml and aliquots can be kept for 1 year at -20°C. Please note that we cannot guarantee the stability of the product if aliquots are made at less than 10 mg/ml. Importantly, avoid repeated freeze-thaw cycles.

What is the selection concentration of Blasticidin in mammalian cells?
The working concentration of Blasticidin for mammalian cell lines varies between 1 - 10 μg/ml, with a few reported cases of up to 30 μg/ml. In an initial experiment we recommend to determine the optimal concentration of Blasticidin required to kill your host cell line.
For more detailed information, please refer to the technical data sheet for this product.

What is the selection concentration of Blasticidin in E.coli?
Blasticidin has low activity in E. coli, but transformants resistant to Blasticidin can be selected on low salt LB agar medium (pH 8) supplemented with 100 μg/ml Blasticidin.
Please note: a high pH (i.e. 8) enhances the activity of Blasticidin.

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