article · 26/06/2019
rEXPAR: An Isothermal Amplification Scheme That Is Robust to Autocatalytic Parasites
Résumé
In the absence of DNA, a solution containing the four deoxynucleotidetriphosphates (dNTPs), a DNA polymerase, and a nicking enzyme generates a self-replicating mixture of DNA species called parasite. Parasites are problematic in template-based isothermal amplification schemes such as EXPAR as well as in related molecular programming approaches, such as the PEN DNA toolbox. Here we show that using a nicking enzyme with only three letters (C, G, T) in the top strand of its recognition site, such as Nb.BssSI, allows us to change the sequence design of EXPAR templates in a way that prevents the formation of parasites when dATP is removed from the solution. This method allows us to make the EXPAR reaction robust to parasite contamination, a common feature in the laboratory, while keeping it compatible with PEN programs, which we demonstrate by engineering a parasite-proof bistable reaction network.
Citer cet article
Urtel, G., Van Der Hofstadt Serrano, M., Galas, J.-C., & Estevez-Torres, A. (2019). rEXPAR: An Isothermal Amplification Scheme That Is Robust to Autocatalytic Parasites. Biochemistry, 58(23), 2675-2681. https://doi.org/10.1021/acs.biochem.9b00063@article{Urtel2019_321,
author = {Urtel, Georg and Van Der Hofstadt Serrano, Marc and Galas, Jean-Christophe and Estevez-Torres, André},
year = {2019},
month = {6},
title = {rEXPAR: An Isothermal Amplification Scheme That Is Robust to Autocatalytic Parasites},
journal = {Biochemistry},
volume = {58},
number = {23},
pages = {2675-2681},
abstract = {In the absence of DNA, a solution containing the four deoxynucleotidetriphosphates (dNTPs), a DNA polymerase, and a nicking enzyme generates a self-replicating mixture of DNA species called parasite. Parasites are problematic in template-based isothermal amplification schemes such as EXPAR as well as in related molecular programming approaches, such as the PEN DNA toolbox. Here we show that using a nicking enzyme with only three letters (C, G, T) in the top strand of its recognition site, such as Nb.BssSI, allows us to change the sequence design of EXPAR templates in a way that prevents the formation of parasites when dATP is removed from the solution. This method allows us to make the EXPAR reaction robust to parasite contamination, a common feature in the laboratory, while keeping it compatible with PEN programs, which we demonstrate by engineering a parasite-proof bistable reaction network.},
url = {https://pubs.acs.org/doi/10.1021/acs.biochem.9b00063},
doi = {10.1021/acs.biochem.9b00063},
}TY - JOUR
AU - Urtel, Georg
AU - Van Der Hofstadt Serrano, Marc
AU - Galas, Jean-Christophe
AU - Estevez-Torres, André
PY - 2019
DA - 2019/06/26
TI - rEXPAR: An Isothermal Amplification Scheme That Is Robust to Autocatalytic Parasites
JO - Biochemistry
VL - 58
IS - 23
AB - In the absence of DNA, a solution containing the four deoxynucleotidetriphosphates (dNTPs), a DNA polymerase, and a nicking enzyme generates a self-replicating mixture of DNA species called parasite. Parasites are problematic in template-based isothermal amplification schemes such as EXPAR as well as in related molecular programming approaches, such as the PEN DNA toolbox. Here we show that using a nicking enzyme with only three letters (C, G, T) in the top strand of its recognition site, such as Nb.BssSI, allows us to change the sequence design of EXPAR templates in a way that prevents the formation of parasites when dATP is removed from the solution. This method allows us to make the EXPAR reaction robust to parasite contamination, a common feature in the laboratory, while keeping it compatible with PEN programs, which we demonstrate by engineering a parasite-proof bistable reaction network.
SP - 2675
EP - 2681
DO - 10.1021/acs.biochem.9b00063
UR - https://pubs.acs.org/doi/10.1021/acs.biochem.9b00063
ER -