TY - GEN
T1 - Improvement of microtas for directed evolution of NAD(P)-dependent oxidoreductases
AU - Goto, Haruna
AU - Kanai, Yuki
AU - Yotsui, Arisa
AU - Shimokihara, Shota
AU - Shitara, Shunya
AU - Einaga, Yasuaki
AU - Matsumoto, Yoshinori
AU - Miki, Norihisa
AU - Fujiwara, Kei
AU - Doi, Nobuhide
N1 - Funding Information:
We thank members of our laboratory for experimental advice and useful discussion. This work was supported by a Grant-in-Aid for Scientific Research (24656508) from the JSPS of Japan, a Keio Gijuku Academic Development Fund, and the Asahi Glass Foundation.
Publisher Copyright:
Copyright © (2018) by Chemical and Biological Microsystems Society. All rights reserved.
PY - 2018
Y1 - 2018
N2 - Here we report an improved μTAS based on electrochemical measurements and microbeads sorting for screening of NAD(P)-dependent oxidoreductases. By using this system, we measured the activity of model enzymes immobilized on microbeads by detection of NADH with a boron-doped diamond (BDD) electrode in each compartment (∼30 nL) of the microfluidic system. Then, we recovered the magnetic microbeads or the droplets containing genes of enzymes with higher activity by magnetophoretic or dielectrophoretic sorting, respectively. Our μTAS should be useful for directed evolution of various NAD(P)-dependent oxidoreductases with high activity and stability in the future.
AB - Here we report an improved μTAS based on electrochemical measurements and microbeads sorting for screening of NAD(P)-dependent oxidoreductases. By using this system, we measured the activity of model enzymes immobilized on microbeads by detection of NADH with a boron-doped diamond (BDD) electrode in each compartment (∼30 nL) of the microfluidic system. Then, we recovered the magnetic microbeads or the droplets containing genes of enzymes with higher activity by magnetophoretic or dielectrophoretic sorting, respectively. Our μTAS should be useful for directed evolution of various NAD(P)-dependent oxidoreductases with high activity and stability in the future.
KW - Boron-doped diamond electrode
KW - Cell-free protein synthesis
KW - Dielectrophoresis
KW - Directed evolution
KW - High-throughput screening
UR - http://www.scopus.com/inward/record.url?scp=85079751775&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85079751775&partnerID=8YFLogxK
M3 - Conference contribution
AN - SCOPUS:85079751775
T3 - 22nd International Conference on Miniaturized Systems for Chemistry and Life Sciences, MicroTAS 2018
SP - 2247
EP - 2249
BT - 22nd International Conference on Miniaturized Systems for Chemistry and Life Sciences, MicroTAS 2018
PB - Chemical and Biological Microsystems Society
T2 - 22nd International Conference on Miniaturized Systems for Chemistry and Life Sciences, MicroTAS 2018
Y2 - 11 November 2018 through 15 November 2018
ER -