Microbiological analysis of 5-formyltetrahydrofolic acid and other folates using an automatic 96-well plate reader☆
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Synthetic biology-driven microbial production of folates: Advances and perspectives
2021, Bioresource TechnologyCitation Excerpt :Accordingly, the efficient mutation library provided by the Himar1 transposon could provide the key basis for identifying 5-MTHF. In addition, the mutant libraries can be screened by high-throughput detection methods based on the growth curve changes of Lactobacillus casei and 5-formyltetrahydrofolate in 96-well plates (Newman and Tsai, 1986). The transposon mutation sites ultimately identified as having an effect on 5-formyltetrahydrofolate production were determined to be folate synthesis pathway genes, and their relevant functions were further verified by knockout and overexpression, as well as by liquid chromatography-mass spectrometry analysis of changes in the ratios of different folates to validate their application for guiding the precise synthesis of 5-MTHF (Fig. 3c).
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2009, HEALTHGRAIN Methods: Analysis of Bioactive Components in Small Grain CerealsAdaptation of Lactobacillus rhamnosus riboflavin assay to microtiter plates
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2006, Analytical BiochemistryCitation Excerpt :The difference in response to 5-CH3-THF observed between the method of Molloy and Scott [132] and the methods of other studies [131,133,134] may be due to the specifics of the conditions employed but points to the need to validate each method with respect to folate species that may be present. Microbiological assays can be highly sensitive [71,126,127] and relatively inexpensive; microtiter plate methods use less than 0.2 ml of media per sample, cryopreserved microbial stocks are cheap and easy to maintain, and determination of growth requires only a microtiter plate reader. In most cases, the assay can be semiautomated using robotic processes.
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This work was supported in part by Grant NIH BRSG S07RR-05471 to the City of Hope National Medical Center. E. M. Newman was supported in part by the Sam Revel Research Fellowship. J. F. Tsai was a student under the Committee for Advanced Science Training program of the California State Museum of Science and Industry.