Blasticidin S Hydrochloride Solution (10 mg/mL in 20mM HEPES)

- Overview
  - Blasticidin S Hydrochloride Solution (10 mg/mL in 20mM HEPES) is a sterile solution of Blasticidin S HCl in 20mM HEPES buffer.
    B006 (10 x 1 mL) contains 10 mg Blasticidin S HCl per vial (100 mg total).
    B007 (20 mL) contains 200 mg Blasticidin S HCl per vial.
    Blasticidin is a peptidyl nucleoside produced by several species of Streptomyces that was first isolated from S. griseochromogenes in 1958. Blasticidin S inhibits protein synthesis and is active against bacteria, fungi, nematodes, and tumor cells. The compound is used as a selection antibiotic for both eukaryotic and prokaryotic cells, and a marker for strain manipulation. It can also be used in genome editing using the CRISPR/Cas9 system.
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    Background
    
    Mechanism of Action
    Blasticidin S Hydrochloride Solution inhibits protein synthesis in prokaryotic and eukaryotic cells by binding to the ribosomal P-site which strengthens tRNA binding and slows down and prevents subsequent peptide synthesis.
    
    Mechanisms of Resistance
    Resistance to Blasticidin S is conferred by bsr, BSD, and bls resistance genes isolated from Bacillus cereus K55-S1, Aspergillus terreus, and Streptoverticillum spp, respectively.
    The bsr resistance gene is a 420 bp fragment and encodes a 15 kDa Blasticidin S deaminase which catalyzes the reaction of Blasticidin S to deaminohydroxyblasticidin S. Deaminohydroxyblasticidin S is a biologically inactive derivative of Blasticidin S and does not interact with or inhibit prokaryotic or eukaryotic ribosomes.
    The BSD resistance gene is a 393 bp fragment and also encodes a Blasticidin S deaminase enzyme which catalyzes a similar reaction to the BSR deaminase. A study by Kimura et al. found the transfection frequency with BSD to be 80X greater than with bsr when using FM3A cells.
    The bls resistance gene encodes an acetyltransferase which interacts with acetyl-coenzyme A and prevents Blasticidin S from inhibiting protein synthesis.
- Properties
  - CAS Number
    
    3513-03-9
    
    Molecular Formula
    
    C17H26N8O5 · HCl
    
    Molecular Weight
    
    458.90 g/mol
    
    Appearance
    
    Clear and colorless or light yellow solution
    
    Other Properties
    
    Source: Streptomyces griseochromogenes
    PH: 7.2-7.5
    Concentration: 10.0±0.3 mg/mL
    Purity Level: Not less than 98.0% (powder, HPLC)
    
    Storage
    
    -20°C
    
    * For research use only
- Applications
  - Application Description
    
    Microbiology Applications: Blasticidin S Hydrochloride Solution can be used as a selection agent after transformation of prokaryotic (bacterial) cells, namely E. coli. Optimal Blasticidin S HCl selection concentrations range from 25 - 100 µg/mL and should be tested for each experimental condition. Selective media containing Blasticidin S HCl should contain a low salt concentration (<90mM) and pH ≤7 to avoid Blasticidin degradation.
    
    Eukaryotic Cell Culture Applications: Blasticidin is a selection antibiotic for both eukaryotic and prokaryotic cells. Resistance to Blasticidin is conferred by the following genes:
    1) bsr (Blasticidin resistance), from Bacillus aureus, which codes for blasticidin-S deaminase.
    2) bls (Blasticidin S acetyltransferase), from Streptoverticillum spp.
    3) BSD (Blasticidin S deaminase), from Apergillus terreus.
    Researchers used Blasticin S (TOKU-E) to select for transfected AS-B145 and BT-474 cells, which are human breast cancer stem/progenitor cells, a subpopulation of cancer cells that are involved in tumor initiation, resistance to therapy, and metastasis (Lu et al, 2016). Typically, mammalian cells are sensitive to Blasticidin S HCl concentrations of 1-10 µg/ml, and bacteria to 25-100 µg/ml. Optimal selection concentration depends on the cell line, growth conditions, media, reagent quality and potency, manufacturing lot, cell density, cell metabolic rate, cell cycle phase, and plasmid properties. A kill curve should be performed for each experimental system to determine the optimal working concentration.
    For more information on relevant cell lines, culture medium, and working concentrations, please visit the TOKU-E Cell-culture Database.