| RRC ID |
59084
|
| 著者 |
Goto H, Kanai Y, Yotsui A, Shimokihara S, Shitara S, Oyobiki R, Fujiwara K, Watanabe T, Einaga Y, Matsumoto Y, Miki N, Doi N.
|
| タイトル |
Microfluidic screening system based on boron-doped diamond electrodes and dielectrophoretic sorting for directed evolution of NAD(P)-dependent oxidoreductases.
|
| ジャーナル |
Lab Chip
|
| Abstract |
We report the development of a micro total analysis system (μTAS) based on electrochemical measurements and dielectrophoretic sorting for screening of NAD(P)-dependent oxidoreductases. In this system, the activity of enzymes immobilized on microbeads, together with their encoding DNA, can be measured with a boron-doped diamond (BDD) electrode in each compartment (∼30 nL) of the microfluidic system. The 30 nL droplets containing microbead-displayed genes of enzymes with higher activity can then be recovered by dielectrophoretic sorting. Previously, we developed the NAD(P)H-measuring device containing the BDD electrode for high-throughput measurement of the activity of NAD(P)-dependent oxidoreductases. In this study, we fabricated an encapsulating device and a droplet-sorting device for nanoliter-size droplets, for the first time, and then combined these three devices to construct a μTAS for directed evolution of NAD(P)-dependent oxidoreductases. We confirmed that this system works by proof-of-principle experiments and successfully applied this system for screening of randomized libraries of NAD-dependent dehydrogenases.
|
| 巻・号 |
20(4)
|
| ページ |
852-861
|
| 公開日 |
2020-2-21
|
| DOI |
10.1039/c9lc01263j
|
| PMID |
31984406
|
| MeSH |
Boron*
Diamond*
Electrodes
Microfluidics
NAD
Oxidoreductases / genetics
|
| IF |
6.914
|
| 引用数 |
0
|
| リソース情報 |
| 原核生物(大腸菌) |
iVEC strain |
