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| IPTG Genomic Grade Powder (Dioxane Free) | |||||||||||||||||||||||||||||||||||||
Properties and Applications: IPTG (isopropylthio-beta-D-thiogalactopyranoside) is an inducer of beta-galactosidase production in bacteria. IPTG is used together with the chromogenic substrates X-gal or Bluo-Gal to determine lac gene expression in recombinant methods. IPTG is a chemical analog of galactose which cannot be cleaved by the enzyme β-galactosidase. Therefore, it can serve as an inducer for activity of the E. coli lac operon by binding and inactivating the lac repressor. A stock solution, 0.1M, is prepared by dissolving IPTG in water with subsequent sterile fi ltration of the solution. The fi nal concentration of IPTG in indicator plates-added after autoclaving should be approximately 0.2mM/L. Analytical Specifi cations: Recommended storage condition: frozen, Molecular weight: 238.21, Appearance: white, free- fl owing powder, Melting point: 118°C
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HyperINDUCE HyperINDUCE™ is an innovative alternative to IPTG that increases the level of soluble, heterologously expressed protein in E. coli containing the T7lac promoter. By decreasing the rate of expression, HyperINDUCE™ enables expressed protein to fold properly and remain soluble thereby reducing the formation of insoluble inclusion bodies. The simplest method for improving yield of soluble protein in E. coli is to reduce the rate of expression. Altering incubation temperature and IPTG concentrations helps, but only marginally. A better alternative for reducing expression rates is to use HyperINDUCE™ with its lower affi nity for the lacI repressor compared with IPTG. Therefore the rates of expression are slower with HyperINDUCE™ resulting in an increased level of soluble protein. Working Concentration: The HyperINDUCE™ is provided at a concentration of 2 mg/mL and a 10X concentrate. A concentration that favors the formation of soluble protein must be determined experimentally, and usually ranges from 2 to 6 g/L. Determining HyperINDUCE™ concentration for optimal protein solubility Pick individual bacteria colonies into 4-6 mL LB or NYZ medium plus the appropriate antibiotic. Incubate in a shaker
(250 RPM) at 37°C overnight or until the culture obtains an O.D.= 0.6-0.8 at 600 nm. In the morning, add 2-3 mL of the
starter culture to 600 ml of medium containing the same antibiotic. Incubate the culture at 37°C until an O.D.= 0.6-0.8
is obtained. Cool the culture on ice to between 15-20°C then split the volume equally into three fl asks. Add the Hyper-
INDUCE™ to a fi nal concentration ranging from 1.0 - 6.0 g/L. Also add the same amount of antibiotic to the culture that
was used initially. Incubate in a shaker (RPM 125) at room temperature for at least 12 hours or overnight. After incubation,
lyse cells and isolate the soluble and insoluble fractions according to standard protocols. Determine the amount
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