Part:BBa_K912025:Experience

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Applications of BBa_K912025

Northwestern 2012 iGEM Team: To obtain a relative activity of the phytase produced by the cells with the constitutive promoter/citrobacter phytase part, we performed a phosphate assay on the produced phytase. In order to remove the phytase from the cells, the cells were overnighted and diluted 1:500 and then regrown into the end of log phase. Once at the end of log phase, the cells were centrifuged and sonicated in order to obtain the intracellular protein(with the produced phytase)to use in a phosphate assay. To perform the phosphate assay, .1 nmol phytic acid was added to the sonicated cell lysate at both a pH of 7 and a pH of 4.5. The pH of 4.5 was the literature value of pH that citrobacter phytase works best at. A colormetric phosphate assay solution was added and the OD of the solution was recorded in comparison to phosphate standards in order to obtain a measure of free phosphate in the solution.

[http://i.imgur.com/qdIsd.png Phosphate_assay_results]

From the above phosphate assay results, we concluded that the produced Citrobacter phytase grown in E. coli managed to liberate phosphate from the phytic acid solution. We are continuing work on performing more phosphate assays to obtain results with more statistical rigor.

Please refer to the results section of our wiki for more information on both this part and related parts http://2012.igem.org/Team:Northwestern/Project/Results

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