Development of a high-throughput screening method for recombinant Escherichia coli with intracellular dextransucrase activity

被引:0
作者
So-Ra Lee
Ah-Rum Yi
Hong-Gyun Lee
Myoung-Uoon Jang
Jung-Mi Park
Nam Soo Han
Tae-Jip Kim
机构
[1] Chungbuk National University,Department of Food Science and Technology
来源
The Journal of Microbiology | 2011年 / 49卷
关键词
high-throughput screening; D-cycloserine; dextransucrase; recombinant ;
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学科分类号
摘要
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.
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