TY - JOUR
T1 - Rapid cell-free forward engineering of novel genetic ring oscillators
AU - Niederholtmeyer, Henrike
AU - Sun, Zachary Z.
AU - Hori, Yutaka
AU - Yeung, Enoch
AU - Verpoorte, Amanda
AU - Murray, Richard M.
AU - Maerkl, Sebastian J.
N1 - Publisher Copyright:
© Niederholtmeyer et al.
PY - 2015/10/2
Y1 - 2015/10/2
N2 - While complex dynamic biological networks control gene expression in all living organisms, the forward engineering of comparable synthetic networks remains challenging. The current paradigm of characterizing synthetic networks in cells results in lengthy design-build-test cycles, minimal data collection, and poor quantitative characterization. Cell-free systems are appealing alternative environments, but it remains questionable whether biological networks behave similarly in cell-free systems and in cells. We characterized in a cell-free system the ‘repressilator’, a three-node synthetic oscillator. We then engineered novel three, four, and five-gene ring architectures, from characterization of circuit components to rapid analysis of complete networks. When implemented in cells, our novel 3-node networks produced population-wide oscillations and 95% of 5-node oscillator cells oscillated for up to 72 hr. Oscillation periods in cells matched the cell-free system results for all networks tested. An alternate forward engineering paradigm using cell-free systems can thus accurately capture cellular behavior.
AB - While complex dynamic biological networks control gene expression in all living organisms, the forward engineering of comparable synthetic networks remains challenging. The current paradigm of characterizing synthetic networks in cells results in lengthy design-build-test cycles, minimal data collection, and poor quantitative characterization. Cell-free systems are appealing alternative environments, but it remains questionable whether biological networks behave similarly in cell-free systems and in cells. We characterized in a cell-free system the ‘repressilator’, a three-node synthetic oscillator. We then engineered novel three, four, and five-gene ring architectures, from characterization of circuit components to rapid analysis of complete networks. When implemented in cells, our novel 3-node networks produced population-wide oscillations and 95% of 5-node oscillator cells oscillated for up to 72 hr. Oscillation periods in cells matched the cell-free system results for all networks tested. An alternate forward engineering paradigm using cell-free systems can thus accurately capture cellular behavior.
UR - http://www.scopus.com/inward/record.url?scp=84949895685&partnerID=8YFLogxK
U2 - 10.7554/eLife.09771
DO - 10.7554/eLife.09771
M3 - Article
C2 - 26430766
AN - SCOPUS:84949895685
SN - 2050-084X
VL - 4
JO - eLife
JF - eLife
IS - OCTOBER2015
M1 - e09771
ER -