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. Author manuscript; available in PMC: 2014 Mar 15.
Published in final edited form as: Methods. 2012 Mar 23;60(1):81–90. doi: 10.1016/j.ymeth.2012.03.009

Figure 1.

Figure 1

Identification of PhlD mutants with improved phloroglucinol production. A PhlD mutant library was constructed through synthetic shuffling. After transformation of the library, membrane-based pre-screening was used to rapidly isolate active clones in a high throughput manner. All active mutants were then picked directly from the membrane into 96-well plates for more quantitative analysis. Phloroglucinol production in cell-free supernatant was quantified using the colorimetric reaction between cinnamaldehyde and phloroglucinol. After characterization of positive mutants, saturation mutagenesis was carried out to evaluate the effect of individual mutations.