A rapid enzymatic assay for high-throughput screening of adenosine-producing strains
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作者:
Dong, Huina
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Chinese Acad Sci, Tianjin Inst Ind Biotechnol, Tianjin 300308, Peoples R China
Chinese Acad Sci, Key Lab Syst Microbial Biotechnol, Tianjin 300308, Peoples R ChinaChinese Acad Sci, Tianjin Inst Ind Biotechnol, Tianjin 300308, Peoples R China
Dong, Huina
[1
,2
]
Zu, Xin
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机构:
Chinese Acad Sci, Tianjin Inst Ind Biotechnol, Tianjin 300308, Peoples R China
Dalian Polytech Univ, Sch Biol Engn, Light Ind Technol & Engn, Dalian 116034, Liaoning, Peoples R ChinaChinese Acad Sci, Tianjin Inst Ind Biotechnol, Tianjin 300308, Peoples R China
Zu, Xin
[1
,3
]
Zheng, Ping
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Chinese Acad Sci, Tianjin Inst Ind Biotechnol, Tianjin 300308, Peoples R China
Chinese Acad Sci, Key Lab Syst Microbial Biotechnol, Tianjin 300308, Peoples R ChinaChinese Acad Sci, Tianjin Inst Ind Biotechnol, Tianjin 300308, Peoples R China
Zheng, Ping
[1
,2
]
Zhang, Dawei
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Chinese Acad Sci, Tianjin Inst Ind Biotechnol, Tianjin 300308, Peoples R China
Chinese Acad Sci, Key Lab Syst Microbial Biotechnol, Tianjin 300308, Peoples R ChinaChinese Acad Sci, Tianjin Inst Ind Biotechnol, Tianjin 300308, Peoples R China
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
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.
机构:
US EPA, Oak Ridge Inst Sci Educ, Res Triangle Pk, NC 27711 USA
US EPA, Integrated Syst Toxicol Div, Res Triangle Pk, NC 27711 USAUS EPA, Oak Ridge Inst Sci Educ, Res Triangle Pk, NC 27711 USA
Paul, Katie B.
Hedge, Joan M.
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US EPA, Integrated Syst Toxicol Div, Res Triangle Pk, NC 27711 USAUS EPA, Oak Ridge Inst Sci Educ, Res Triangle Pk, NC 27711 USA
Hedge, Joan M.
Rotroff, Daniel M.
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US EPA, Natl Ctr Computat Toxicol, Off Res & Dev, Res Triangle Pk, NC 27711 USAUS EPA, Oak Ridge Inst Sci Educ, Res Triangle Pk, NC 27711 USA
Rotroff, Daniel M.
Hornung, Michael W.
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US EPA, Midcontinent Ecol Div, Natl Hlth & Environm Effects Res Lab, Res Triangle Pk, NC 27711 USAUS EPA, Oak Ridge Inst Sci Educ, Res Triangle Pk, NC 27711 USA
Hornung, Michael W.
Crofton, Kevin M.
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US EPA, Natl Ctr Computat Toxicol, Off Res & Dev, Res Triangle Pk, NC 27711 USAUS EPA, Oak Ridge Inst Sci Educ, Res Triangle Pk, NC 27711 USA
Crofton, Kevin M.
Simmons, Steven O.
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US EPA, Integrated Syst Toxicol Div, Res Triangle Pk, NC 27711 USAUS EPA, Oak Ridge Inst Sci Educ, Res Triangle Pk, NC 27711 USA
机构:
Univ N Carolina, Sch Med, Dept Biochem & Biophys, Chapel Hill, NC 27599 USAUniv N Carolina, UNC Eshelman Sch Pharm, Div Chem Biol & Med Chem, Chapel Hill, NC 27599 USA
Peterson, Eliza J. R.
Janzen, William P.
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Univ N Carolina, UNC Eshelman Sch Pharm, Div Chem Biol & Med Chem, Chapel Hill, NC 27599 USA
Univ N Carolina, UNC Eshelman Sch Pharm, Ctr Integrat Chem Biol & Drug Discovery, Chapel Hill, NC 27599 USAUniv N Carolina, UNC Eshelman Sch Pharm, Div Chem Biol & Med Chem, Chapel Hill, NC 27599 USA
Janzen, William P.
Kireev, Dmitri
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Univ N Carolina, UNC Eshelman Sch Pharm, Div Chem Biol & Med Chem, Chapel Hill, NC 27599 USA
Univ N Carolina, UNC Eshelman Sch Pharm, Ctr Integrat Chem Biol & Drug Discovery, Chapel Hill, NC 27599 USAUniv N Carolina, UNC Eshelman Sch Pharm, Div Chem Biol & Med Chem, Chapel Hill, NC 27599 USA
Kireev, Dmitri
Singleton, Scott F.
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Univ N Carolina, UNC Eshelman Sch Pharm, Div Chem Biol & Med Chem, Chapel Hill, NC 27599 USA
Univ N Carolina, Sch Med, Dept Biochem & Biophys, Chapel Hill, NC 27599 USAUniv N Carolina, UNC Eshelman Sch Pharm, Div Chem Biol & Med Chem, Chapel Hill, NC 27599 USA