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S-Gal is a trademark product of Sigma Aldrich for the compound 3,4-cyclohexenoesculetin-
S-Gal is a trademark product of Sigma Aldrich for the compound 3,4-cyclohexenoesculetin-
β-D-galactopyranoside
β-D-galactopyranoside, C19H21O9Na, MW 416.35, MDL Number MFCD03458481.
S-Gal®, sodium salt is a patented water-soluble, autoclavable chromogenic substrate for β-galactosidase that is designed to replace X-Gal in blue-white selection of recombinant bacterial colonies with the lac+ phenotype. Caution: For black color development to occur, ferric ion must be present. Although we recommend ferric ammonium citrate (500 mg/L of media), other ferric compounds can be used to provide this requirement, depending upon your particular system. From [http://www.sigmaaldrich.com/catalog/ProductDetail.do?N4=S7313%7CSIGMA&N5=SEARCH_CONCAT_PNO%7CBRAND_KEY&F=SPEC Sigma-Aldrich site].
[[Image:sgal.tiff|400px|left]]
[[Image:sgal.tiff|400px|left]]
Image from Heuermann and Cosgrove PMID 11355350
Image from Heuermann and Cosgrove PMID 11355350
Reagents ordered in February 2009
Reagents ordered in February 2009
0F5879-100G, $35.54, AMMONIUM IRON(III) CITRATE, REAGENT GRADE
0F5879-100G, $35.54, AMMONIUM IRON(III) CITRATE, REAGENT GRADE
S7313-1G, $385.60, S-GAL(R) SODIUM SALT
S7313-1G, $385.60, S-GAL(R) SODIUM SALT
==S-Gal staining conditions==
==S-Gal staining conditions==
According to [http://www.sigmaaldrich.com/catalog/ProductDetail.do?N4=S7313%7CSIGMA&N5=SEARCH_CONCAT_PNO%7CBRAND_KEY&F=SPEC Sigma-Aldrich site]:
Stock solutions of S-Gal can be made by dissolving at 50 mg/mL in deionized water, sterile-filtering and storing at -20 °C.
'''For detecting plasmids on agar plates, from Heuermann and Cosgrove PMID 11355350'''
'''For detecting plasmids on agar plates, from Heuermann and Cosgrove PMID 11355350'''
For chemical analysis, ferric ammonium citrate, S-Gal, and IPTG were added to 50-mL aliquots LB broth or deionized water at final concentrations of 500, 300, and 30 µg/mL, respectively (all from Sigma). S-Gal dye was initially dissolved in N,N-dimethylformamide (DMF) (Sigma) at a concentration of 200 mg/mL and was added to water or medium for analysis at the concentration stated above before autoclaving or microwaving. For this part of the evaluation, a stock solution of S-Gal in DMF was used for convenience and for the consistent addition of S-Gal to media or aqueous solu- tions. S-Gal, determined empirically to be approximately 70% more soluble than X-gal at room temperature (data not shown), is suitable for the blended-medium product but re- quires dissolving in DMF for the preparation of more concen- trated solutions such as the stock solution described here. IPTG was added to the water or medium from an aqueous stock of 100 mg/mL. Ferric ammonium citrate was prepared as an aqueous stock solution of 200 mg/mL.
For chemical analysis, ferric ammonium citrate, S-Gal, and IPTG were added to 50-mL aliquots LB broth or deionized water at final concentrations of 500, 300, and 30 µg/mL, respectively (all from Sigma). S-Gal dye was initially dissolved in N,N-dimethylformamide (DMF) (Sigma) at a concentration of 200 mg/mL and was added to water or medium for analysis at the concentration stated above before autoclaving or microwaving. For this part of the evaluation, a stock solution of S-Gal in DMF was used for convenience and for the consistent addition of S-Gal to media or aqueous solutions. S-Gal, determined empirically to be approximately 70% more soluble than X-gal at room temperature (data not shown), is suitable for the blended-medium product but requires dissolving in DMF for the preparation of more concen- trated solutions such as the stock solution described here. IPTG was added to the water or medium from an aqueous stock of 100 mg/mL. Ferric ammonium citrate was prepared as an aqueous stock solution of 200 mg/mL.
S-Gal, IPTG and ferric ammonium citrate are dry-blended with standard LB agar at the final concentrations stated above. The
S-Gal, IPTG and ferric ammonium citrate are dry-blended with standard LB agar at the final concentrations stated above. The powdered blend was suspended in deionized water and autoclaved as previously described. For the preparation of microwaved medium, the suspended blend was heated until boiling, followed by swirling to allow the remaining agar component to go into solution. In this case, the antibiotic ampicillin was added after autoclaving (100 mg/mL).
powdered blend was suspended in deionized water and autoclaved as previously described. For the preparation of microwaved medium, the suspended blend was heated until boiling, followed by swirling to allow the remaining agar component to go into solution. In this case, the antibiotic ampicillin was added after autoclaving (100 mg/mL).