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Figure 1

Nature Methods 3, 609 - 614 (2006)
Published online: 21 July 2006; | doi:10.1038/nmeth899

High-throughput screening methodology for the directed evolution of glycosyltransferases

Amir Aharoni, Karena Thieme, Cecilia P C Chiu, Sabrina Buchini, Luke L Lairson, Hongming Chen, Natalie C J Strynadka, Warren W Wakarchuk & Stephen G Withers

 
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Figure 1. Cell-based assay for ST activity.
A gene library is transformed and cloned in E. coli (1). The encoded ST protein is expressed in the cytoplasm of the engineered JM107 NanA- strain together with CMP-Neu5Ac synthetase (2). Cells are incubated with Neu5Ac donor sugar and fluorescently labeled acceptor sugars (3). After incubation, cells are extensively washed to remove unreacted fluorescent acceptor sugar (4). Cells are directly analyzed and sorted by FACS (5).

 
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