| Title |
NOTE] Development of a High-Throughput Screening Method for Recombinant Escherichia coli with Intracellular Dextransucrase Activity |
| Author |
So-Ra Lee, Ah-Rum Yi, Hong-Gyun Lee, Myoung-Uoon Jang, Jung-Mi Park, Nam Soo Han, and Tae-Jip Kim* |
| Address |
Department of Food Science and Technology, Chungbuk National University, Cheongju 361-763, Republic of Korea |
| Bibliography |
Journal of Microbiology, 49(2),320-323, 2011,
|
| DOI |
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| Key Words |
high-throughput screening, D-cycloserine, dextransucrase, recombinant E. coli |
| Abstract |
To efficiently engineer intracellular dextransucrase (DSase) expression in Escherichia coli, a high-throughput screening method was developed based on the polymer-forming activity of the enzyme. Recombinant E. coli containing the Leuconostoc citreum DSase (LcDS) gene was grown on Luria-Bertani agar plates, containing 2% sucrose, at 37°C for 8 h. The plates were then evenly overlaid with 0.6% soft agar, containing 1.2 mg/ml D-cycloserine, and incubated at 30°C to allow gradual cell disruption until a dextran polymer grew through the overlaid layer. A significant correlation between dextran size and enzyme activity was established and applied for screening truncated mutants with LcDS activity. |
