A rapid enzymatic assay for high-throughput screening of adenosine-producing strains

被引:6
|
作者
Dong, Huina [1 ,2 ]
Zu, Xin [1 ,3 ]
Zheng, Ping [1 ,2 ]
Zhang, Dawei [1 ,2 ]
机构
[1] Chinese Acad Sci, Tianjin Inst Ind Biotechnol, Tianjin 300308, Peoples R China
[2] Chinese Acad Sci, Key Lab Syst Microbial Biotechnol, Tianjin 300308, Peoples R China
[3] Dalian Polytech Univ, Sch Biol Engn, Light Ind Technol & Engn, Dalian 116034, Liaoning, Peoples R China
关键词
NO-REFLOW PHENOMENON; BACILLUS-SUBTILIS; INTRACORONARY ADENOSINE; REPERFUSION INJURY; AMMONIA; APTASENSOR; SERUM; CHROMATOGRAPHY; ATTENUATION; SYNTHETASE;
D O I
10.1111/1751-7915.12189
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
Adenosine is a major local regulator of tissue function and industrially useful as precursor for the production of medicinal nucleoside substances. High-throughput screening of adenosine overproducers is important for industrial microorganism breeding. An enzymatic assay of adenosine was developed by combined adenosine deaminase (ADA) with indophenol method. The ADA catalyzes the cleavage of adenosine to inosine and NH3, the latter can be accurately determined by indophenol method. The assay system was optimized to deliver a good performance and could tolerate the addition of inorganic salts and many nutrition components to the assay mixtures. Adenosine could be accurately determined by this assay using 96-well microplates. Spike and recovery tests showed that this assay can accurately and reproducibly determine increases in adenosine in fermentation broth without any pretreatment to remove proteins and potentially interfering low-molecular-weight molecules. This assay was also applied to high-throughput screening for high adenosine-producing strains. The high selectivity and accuracy of the ADA assay provides rapid and high-throughput analysis of adenosine in large numbers of samples.
引用
收藏
页码:230 / 238
页数:9
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