PDF(1952 KB)
PDF(1952 KB)
PDF(1952 KB)
一种可实现合成生物传感器现场部署的增材制造方法
An Additive Manufacturing Approach that Enables the Field Deployment of Synthetic Biosensors
The tools of synthetic biology can be used to engineer living biosensors that report the presence of analytes. Although these engineered cellular biosensors have many potential applications for deployment outside of the lab, they are genetically modified organisms (GMOs) and are often considered dangerous. Mitigating the risk of releasing GMOs into the environment while enabling their use outside a laboratory is critical. Here, we describe the development of a biosensing system consisting of a synthetic biological circuit, which is engineered in Escherichia coli that are contained within a unique 3D-printed device housing. These GMOs detect the chemical quorum signal of Pseudomonas aeruginosa, an opportunistic pathogen. Using this device, the living biosensor makes contact with a specimen of interest without ever being exposed to the environment. Cells can be visually analyzed in the field within culture tubes, or returned to the lab for further analysis. Many biosensors lack the versatility required for deployment in the field, where many diseases can go undiagnosed due to a lack of resources and equipment. Our bioassay device utilizes 3D printing to create a portable, modular, and inexpensive device for the field deployment of living biosensors.
Synthetic biology / Additive manufacturing / Biosensors
| [1] |
Gardner T.S., Cantor C.R., Collins J.J.. Construction of a genetic toggle switch in Escherichia coli. Nature. 2000; 403(6767): 339-342.
|
| [2] |
Elowitz M.B., Leibler S.. A synthetic oscillatory network of transcriptional regulators. Nature. 2000; 403(6767): 335-338.
|
| [3] |
Balagaddé F.K., Song H., Ozaki J., Collins C.H., Barnet M., Arnold F.H.,
|
| [4] |
Litcofsky K.D., Afeyan R.B., Krom R.J., Khalil A.S., Collins J.J.. Iterative plug-and-play methodology for constructing and modifying synthetic gene networks. Nat Methods. 2012; 9(11): 1077-1080.
|
| [5] |
Brenner K., Karig D.K., Weiss R., Arnold F.H.. Engineered bidirectional communication mediates a consensus in a microbial biofilm consortium. Proc Natl Acad Sci USA. 2007; 104(44): 17300-17304.
|
| [6] |
Tamsir A., Tabor J.J., Voigt C.A.. Robust multicellular computing using genetically encoded NOR gates and chemical ‘wires’. Nature. 2011; 469(7329): 212-215.
|
| [7] |
McDaniel L.E., Bailey E.G., Zimmerli A.. Effect of oxygen-supply rates on growth of Escherichia coli. I. Studies in unbaffled and baffled shake flasks. Appl Microbiol. 1965; 13: 109-114.
|
| [8] |
Ratkowsky D.A., Olley J., McMeekin T.A., Ball A.. Relationship between temperature and growth rate of bacterial cultures. J Bacteriol. 1982; 149(1): 1-5.
|
| [9] |
Kuzma J., Besley J.C.. Ethics of risk analysis and regulatory review: from bio- to nanotechnology. NanoEthics. 2008; 2(2): 149-162.
|
| [10] |
Gregorowius D., Lindemann-Matthies P., Huppenbauer M.. Ethical discourse on the use of genetically modified crops: a review of academic publications in the fields of ecology and environmental ethics. J Agric Environ Ethics. 2012; 25(3): 265-293.
|
| [11] |
Kilama W.L.. Health research ethics in public health: trials and implementation of malaria mosquito control strategies. Acta Trop. 2009; 112(Suppl 1): S37-S47.
|
| [12] |
Schmidt M.. Special issue: societal aspects of synthetic biology. Syst Synth Biol. 2009; 3(1–4): 1-2.
|
| [13] |
Kuzma J., Tanji T.. Unpackaging synthetic biology: identification of oversight policy problems and options. Regul Governance. 2010; 4(1): 92-112.
|
| [14] |
Melchels F.P.W., Feijen J., Grijpma D.W.. A review on stereolithography and its applications in biomedical engineering. Biomaterials. 2010; 31(24): 6121-6130.
|
| [15] |
Dimitrov D., Schreve K., De Beer N.. Advances in three dimensional printing—state of the art and future perspectives. Rapid Prototyping J. 2006; 12(3): 136-147.
|
| [16] |
Schubert C., Van Langeveld M.C., Donoso L.A.. Innovations in 3D printing: a 3D overview from optics to organs. Br J Ophthalmol. 2014; 98(2): 159-161.
|
| [17] |
Ventola C.L.. Medical applications for 3D printing: current and projected uses. PT. 2014; 39(10): 704-711.
|
| [18] |
Gambello M.J., Iglewski B.H.. Cloning and characterization of the Pseudomonas aeruginosa lasR gene, a transcriptional activator of elastase expression. J Bacteriol. 1991; 173(9): 3000-3009.
|
| [19] |
Kiratisin P., Tucker K.D., Passador L.. LasR, a transcriptional activator of Pseudomonas aeruginosa virulence genes, functions as a multimer. J Bacteriol. 2002; 184(17): 4912-4919.
|
| [20] |
Schwarzer C., Fu Z., Fischer H., Machen T.E.. Redox-independent activation of NF-κB by Pseudomonas aeruginosa pyocyanin in a cystic fibrosis airway epithelial cell line. J Biol Chem. 2008; 283(40): 27144-27153.
|
| [21] |
Pearson J.P., Pesci E.C., Iglewski B.H.. Roles of Pseudomonas aeruginosa las and rhl quorum-sensing systems in control of elastase and rhamnolipid biosynthesis genes. J Bacteriol. 1997; 179(18): 5756-5767.
|
| [22] |
Seed P.C., Passador L., Iglewski B.H.. Activation of the Pseudomonas aeruginosa lasI gene by LasR and the Pseudomonas autoinducer PAI: an autoinduction regulatory hierarchy. J Bacteriol. 1995; 177(3): 654-659.
|
| [23] |
Lyczak J.B., Cannon C.L., Pier G.B.. Lung infections associated with cystic fibrosis. Clin Microbiol Rev. 2002; 15(2): 194-222.
|
| [24] |
Maciá M.D., Blanquer D., Togores B., Sauleda J., Pérez J.L., Oliver A.. Hypermutation is a key factor in development of multiple-antimicrobial resistance in Pseudomonas aeruginosa strains causing chronic lung infections. Antimicrob Agents Chemother. 2005; 49(8): 3382-3386.
|
| [25] |
Pier G.B., Grout M., Zaidi T.S., Olsen J.C., Johnson L.G., Yankaskas J.R.,
|
| [26] |
Pedersen S.S., Espersen F., Høiby N.. Diagnosis of chronic Pseudomonas aeruginosa infection in cystic fibrosis by enzyme-linked immunosorbent assay. J Clin Microbiol. 1987; 25(10): 1830-1836.
|
| [27] |
McCulloch E., Lucas C., Ramage G., Williams C.. Improved early diagnosis of Pseudomonas aeruginosa by real-time PCR to prevent chronic colonisation in a paediatric cystic fibrosis population. J Cyst Fibros. 2011; 10(1): 21-24.
|
| [28] |
Pearson J.P., Gray K.M., Passador L., Tucker K.D., Eberhard A., Iglewski B.H.,
|
| [29] |
Maniatis T., Fritsch E.F., Sambrook J.. Molecular cloning: a laboratory manual. 2nd ed.
|
| [30] |
Ausubel F.M., Brent R., Kingston R., Moore D.D., Seidman J., Struhl K.,
|
| [31] |
Brophy J.A., Voigt C.A.. Principles of genetic circuit design. Nat Methods. 2014; 11(5): 508-520.
|
| [32] |
Hindmarsh A.C.. LSODE and LSODI, two new initial value ordinary differential equation solvers. ACM-SIGNUM Newslett. 1980; 15(4): 10-11.
|
| [33] |
COMSOL. COMSOL multiphysics: user’s guide (version 4.3 a).
|
| [34] |
Szykiedans K., Credo W.. Mechanical properties of FDM and SLA low-cost 3-D prints. Procedia Eng. 2016; 136: 257-262.
|
| [35] |
Pardee K., Green A.A., Ferrante T., Cameron D.E., DaleyKeyser A., Yin P.,
|
| [36] |
Dubin P.J., Kolls J.K.. IL-23 mediates inflammatory responses to mucoid Pseudomonas aeruginosa lung infection in mice. Am J Physiol Lung Cell Mol Physiol. 2007; 292(2): L519-L528.
|
| [37] |
Jensen T., Pedersen S.S., Garne S., Heilmann C., Høiby N., Koch C.. Colistin inhalation therapy in cystic fibrosis patients with chronic Pseudomonas aeruginosa lung infection. J Antimicrob Chemother. 1987; 19(6): 831-838.
|
| [38] |
Ciofu O., Riis B., Pressler T., Poulsen H.E., Høiby N.. Occurrence of hypermutable Pseudomonas aeruginosa in cystic fibrosis patients is associated with the oxidative stress caused by chronic lung inflammation. Antimicrob Agents Chemother. 2005; 49(6): 2276-2282.
|
| [39] |
Heeckeren A., Walenga R., Konstan M.W., Bonfield T., Davis P.B., Ferkol T.. Excessive inflammatory response of cystic fibrosis mice to bronchopulmonary infection with Pseudomonas aeruginosa. J Clin Invest. 1997; 100(11): 2810-2815.
|
| [40] |
Jahoor A., Patel R., Bryan A., Do C., Krier J., Watters C.,
|
| [41] |
Kruger N.J.. The Bradford method for protein quantitation. In:
|
| [42] |
Swanson A.B., Matev I.B., de Groot G.. The strength of the hand. Bull Prosthet Res. 1970; 10(14): 145-153.
|
The authors acknowledge support from funding from federal agencies of the United States including, the National Science Foundation (1709238), as well as funding from Office of Naval Research (N00014-17-12306 and N00014-15-1-2502), and the Air Force Office of Scientific Research (FA9550-13-1-0108). No funding agencies played a significant role in the study design.
Wolozny and Ruder conceived the idea for the system described here. All authors designed and performed the experiments, analyzed the data, discussed the results, wrote this manuscript and commented on the paper. Wolozny, Lake, Long, and Ruder revised the manuscript.
Daniel Wolozny, John R. Lake, Paul G. Movizzo, Zhicheng Long, and Warren C. Ruder declare that they have no conflict of interest or financial conflicts to disclose.
/
| 〈 |
|
〉 |
