Significant research progress in synthetic biology

doi:10.12211/2096-8280.2020-057

Abstract

Abstract:

As an emerging, interdisciplinary field, synthetic biology has made great advances in many directions due to wide acceptance of its core principles and rapid progress in DNA synthesis. In this paper, recent development in gene circuits, genome design and synthesis, cell factories, and synthetic microbial consortia is reviewed. The complexity of artificial gene circuits that can be designed and constructed is gradually increasing with more refined control. Synthetic genomes are routinely assembled, expanding from prokaryotes (Mycoplasma to Escherichia coli) to eukaryotes (Saccharomyces cerevisiae), and improved capacity in genome design promotes the research of biological evolution. Metabolic pathways of ever-increasing lengths are constructed based on modularization and orthogonality principles to produce molecules of complex structures, and fundamental rewiring of cellular metabolism is performed for enhanced robustness and compatibility. The design and construction of synthetic microbial consortia have been expanded from two-species systems to multi-species systems, so that more sophisticated functions can be achieved. At the end of this paper, new research directions resulted from the interdisciplinary integration of synthetic biology and other disciplines are discussed.

Key words: synthetic biology, gene circuit, genome design and synthesis, cell factory, synthetic microbial consortia

摘要：

合成生物学作为一个新兴的交叉学科领域，随着DNA合成技术的进步和合成生物学理念的深入，多个研究方向取得了长足发展。本文主要对基因回路、基因组设计合成、细胞工厂和人工多细胞体系的进展进行了综述。可设计构建的人工基因线路的复杂度逐步提升，人工控制更加精细；组装技术取得快速进展的同时，人工基因组的设计深度也在不断拓展，设计合成的人工基因组由支原体拓展向大肠杆菌，甚至真核生物酿酒酵母，推动了生物进化演化的研究；细胞工厂的设计构建在逐步挑战代谢途径更长、复杂程度更高的化合物的合成，模块化和正交化策略对复杂细胞工厂构建的支撑作用日益明显，鲁棒性和适配性成为细胞工厂构建需要考虑的重要问题；人工多细胞体系的设计构建已经从设计构建两菌体系向多菌体系扩展，通过多种原则进行设计，实现更加复杂的预期功能。本文也对合成生物学与其他学科交叉融合产生的一些新研究方向进行了简介。

关键词: 合成生物学, 基因回路, 基因组设计合成, 细胞工厂, 人工多细胞体系

CLC Number:

Mingzhu DING, Bingzhi LI, Ying WANG, Zexiong XIE, Duo LIU, Yingjin YUAN. Significant research progress in synthetic biology[J]. Synthetic Biology Journal, 2020, 1(1): 7-28.

丁明珠, 李炳志, 王颖, 谢泽雄, 刘夺, 元英进. 合成生物学重要研究方向进展[J]. 合成生物学, 2020, 1(1): 7-28.

Figures/Tables 4

References 238

1	LEPROUST E M, PECK B J, SPIRIN K, et al. Synthesis of high-quality libraries of long (150mer) oligonucleotides by a novel depurination controlled process[J]. Nucleic Acids Research, 2010, 38 (8): 2522-2540.
2	PALLUK S, ARLOW D H, DE ROND T, et al. De novo DNA synthesis using polymerase-nucleotide conjugates[J]. Nature Biotechnology, 2018, 36 (7): 645-650.
3	ELOWITZ M B， LEIBLER S. A synthetic oscillatory network of transcriptional regulators[J]. Nature, 2000, 403 (6767): 335-338.
4	GARDNER T S, CANTOR C R, COLLINS J J. Construction of a genetic toggle switch in Escherichia coli[J]. Nature, 2000, 403 (6767): 339-342.
5	RANTASALO A, KUIVANEN J, PENTTILA M, et al. Synthetic toolkit for complex genetic circuit engineering in Saccharomyces cerevisiae[J]. ACS Synthtic Biology, 2018, 7 (6): 1573-1587.
6	ZONG Y, ZHANG H M, LYU C, et al. Insulated transcriptional elements enable precise design of genetic circuits[J]. Nature Communications, 2017, 8 (1): 52.
7	SHETTY R P, ENDY D， KNIGHT T F JR. Engineering BioBrick vectors from BioBrick parts[J]. J. Biol. Eng., 2008, 2: 5.
8	LEE T S, KRUPA R A, ZHANG F Z, et al. BglBrick vectors and datasheets: a synthetic biology platform for gene expression[J]. Journal of Biological Engineering, 2011, 5 (1): 12.
9	AGMON N, MITCHELL L A, CAI Y Z, et al. Yeast golden gate (yGG) for the efficient assembly of s-cerevisiae transcription Units[J]. ACS Synthetic Biology, 2015, 4 (7): 853-859.
10	WEBER E, ENGLER C, GRUETZNER R, et al. A modular cloning system for standardized assembly of multigene constructs[J]. PLoS One, 2011, 6 (2): e16765.
11	BERVOETS I, BREMPT M VAN, NEROM K VAN, et al. A sigma factor toolbox for orthogonal gene expression in Escherichia coli[J]. Nucleic Acids Research, 2018, 46 (4): 2133-2144.
12	REDDEN H， ALPER H S. The development and characterization of synthetic minimal yeast promoters[J]. Nature Communications, 2015, 6: e7810.
13	ROGERS J K, TAYLOR N D, CHURCH G M. Biosensor-based engineering of biosynthetic pathways[J]. Current Opinion in Biotechnology, 2016, 42: 84-91.
14	TEMME K, HILL R, SEGALL-SHAPIRO T H, et al. Modular control of multiple pathways using engineered orthogonal T7 polymerases[J]. Nucleic Acids Research, 2012, 40 (17): 8773-8781.
15	MORSE N J, WAGNER J M, REED K B, et al. T7 polymerase expression of guide RNAs in vivo allows exportable CRISPR-Cas9 editing in multiple yeast hosts[J]. ACS Synthetic Biology, 2018, 7 (4): 1075-1084.
16	LIAO W, LIU B, CHANG C C, et al. Functional characterization of insulation effect for synthetic gene circuits in mammalian cells[J]. ACS Synthetic Biology, 2018, 7 (2): 412-418.
17	BATEY R T, GILBERT S D, MONTANGE R K. Structure of a natural guanine-responsive riboswitch complexed with the metabolite hypoxanthine[J]. Nature, 2004, 432 (7015): 411-415.
18	BADORREK C S, GHERGHE C M, WEEKS K M. Structure of an RNA switch that enforces stringent retroviral genomic RNA dimerization[J]. PNAS, 2006, 103 (37): 13640-13645.
19	MOON T S, LOU C B, TAMSIR A, et al. Genetic programs constructed from layered logic gates in single cells[J]. Nature, 2012, 491 (7423): 249-253.
20	SIUTI P, YAZBEK J, LU T K. Synthetic circuits integrating logic and memory in living cells[J]. Nature Biotechnology, 2013, 31 (5): 448-452.
21	STRICKER J, COOKSON S, BENNETT M R, et al. A fast, robust and tunable synthetic gene oscillator[J]. Nature, 2008, 456 (7221): 516-519.
22	ANDREWS L B, NIELSEN A A K, VOIGT C A. Cellular checkpoint control using programmable sequential logic[J]. Science, 2018, 361 (6408): eaap8987.
23	LIAO M J, DIN M O, TSINRING L, et al. Rock-paper-scissors: engineered population dynamics increase genetic stability[J]. Science, 2019, 365 (6457): 1045-1049.
24	POTVIN-TROTTIER L, LORD N D, VINNICOMBE G, et al. Synchronous long-term oscillations in a synthetic gene circuit[J]. Nature, 2016, 538 (7626): 514-517.
25	GAO X J, CHONG L S, KIM M S, et al. Programmable protein circuits in living cells[J]. Science, 2018, 361 (6408): 1252-1258.
26	ADAMALA K P, MARTIN-ALARCON D A, GUTHRIE-HONEA K R, et al. Engineering genetic circuit interactions within and between synthetic minimal cells[J]. Nature Chemistry, 2017, 9 (5): 431-439.
27	PURCELL O, WANG J, SIUTI P, et al. Encryption and steganography of synthetic gene circuits[J]. Nature Communications, 2018, 9 (1): 4942.
28	CHEN M T, WEISS R. Artificial cell-cell communication in yeast Saccharomyces cerevisiae using signaling elements from Arabidopsis thaliana[J]. Nature Biotechnology, 2005, 23 (12): 1551-1555.
29	YOU L C, COX R S, WEISS R, et al. Programmed population control by cell-cell communication and regulated killing[J]. Nature, 2004, 428 (6985): 868-871.
30	BALAGADDE F K, SONG H, OZAKI J, et al. A synthetic Escherichia coli predator-prey ecosystem[J]. Molecular Systems Biology, 2008, 4: 187.
31	LIU C L, FU X F, LIU L L, et al. Sequential establishment of stripe patterns in an expanding cell population[J]. Science, 2011, 334 (6053): 238-241.
32	TAMSIR A, TABOR J J, VOIGT C A. Robust multicellular computing using genetically encoded NOR gates and chemical 'wires'[J]. Nature, 2011, 469 (7329): 212-215.
33	REGOT S, MACIA J, CONDE N, et al. Distributed biological computation with multicellular engineered networks[J]. Nature, 2011, 469 (7329): 207-211.
34	TODA S, BLAUCH L R, TANG S K Y, et al. Programming self-organizing multicellular structures with synthetic cell-cell signaling[J]. Science, 2018, 361 (6398): 156-162.
35	ZHANG F Z, CAROTHERS J M, KEASLING J D. Design of a dynamic sensor-regulator system for production of chemicals and fuels derived from fatty acids[J]. Nature Biotechnology, 2012, 30 (4): 354-359.
36	CHOU H H, KEASLING J D. Programming adaptive control to evolve increased metabolite production[J]. Nature Communications, 2013, 4: 2595.
37	TAO R K, ZHAO Y Z, CHU H Y, et al. Genetically encoded fluorescent sensors reveal dynamic regulation of NADPH metabolism[J]. Nature Methods, 2017, 14 (9): 720-728.
38	TAY P K R, NGUYEN P Q, JOSHI N S. A Synthetic circuit for mercury bioremediation using self assembling functional amyloids[J]. ACS Synthetic Biology, 2017, 6 (10): 1841-1850.
39	WAN X Y, VOLPETTI F, PETROVA E, et al. Cascaded amplifying circuits enable ultrasensitive cellular sensors for toxic metals[J]. Nature Chemical Biology, 2019, 15 (5): 540-548.
40	GALLAGHER R R, PATEL J R, INTERIANO A L, et al. Multilayered genetic safeguards limit growth of microorganisms to defined environments[J]. Nucleic Acids Research, 2015, 43 (3): 1945-1954.
41	CHAN C T Y, LEE J W, CAMERON D E, et al. 'Deadman' and 'Passcode' microbial kill switches for bacterial containment[J]. Nature Chemical Biology, 2016, 12 (2): 82-86.
42	ROYBAL K T, RUPP L J, MORSUT L, et al. Precision tumor recognition by T cells with combinatorial antigen-sensing circuits[J]. Cell, 2016, 164 (4): 770-779.
43	ROYBAL K T, WILLIAMS J Z, MORSUT L, et al. Engineering T cells with customized therapeutic response programs using synthetic notch receptors[J]. Cell, 2016, 167 (2): 419-432.
44	MIMEE M, NADEAU P, HAYWARD A, et al. An ingestible bacterial-electronic system to monitor gastrointestinal health[J]. Science, 2018, 360 (6391): 915-918.
45	SHAO J W, XUE S, YU G L, et al. Smartphone-controlled optogenetically engineered cells enable semiautomatic glucose homeostasis in diabetic mice[J]. Science Translational Medicine, 2017, 9 (387): eaal2298.
46	TIGGES M, DENERVAUD N, GREBER D, et al. A synthetic low-frequency mammalian oscillator[J]. Nucleic Acids Research, 2010, 38 (8): 2702-2711.
47	TIGGES M, MARQUEZ-LAGO T T, STELLING J, et al. A tunable synthetic mammalian oscillator[J]. Nature, 2009, 457 (7227): 309-312.
48	GIBSON D G, GLASS J I, LARTIGUE C, et al. Creation of a bacterial cell controlled by a chemically synthesized genome[J]. Science, 2010, 329 (5987): 52-56.
49	HUTCHISON C A, CHUANG R Y, NOSKOV V N, et al. Design and synthesis of a minimal bacterial genome[J]. Science, 2016, 351 (6280): aad6253.
50	OSTROV N, LANDON M, GUELL M, et al. Design, synthesis, and testing toward a 57-codon genome[J]. Science, 2016, 353 (6301): 819-822.
51	FREDENS J, WANG K, DE LA TORRE D, et al. Total synthesis of Escherichia coli with a recoded genome[J]. Nature, 2019, 569 (7757): 514-518.
52	ANNALURU N, MULLER H, MITCHELL L A, et al. Total synthesis of a functional designer eukaryotic chromosome[J]. Science, 2014, 344 (6179): 55-58.
53	XIE Z X, LI B Z, MITCHELL L A, et al. Perfect designer chromosome Ⅴ and behavior of a ring derivative[J]. Science, 2017, 355 (6329): eaaf4704.
54	RICHARDSON S M, MITCHELL L A, STRACQUADANIO G, et al. Design of a synthetic yeast genome[J]. Science, 2017, 355 (6329): 1040-1044.
55	MITCHELL L A, WANG A, STRACQUADANIO G, et al. Synthesis, debugging, and effects of synthetic chromosome consolidation: syn Ⅵ and beyond[J]. Science, 2017, 355 (6329): eaaf4831.
56	SHEN Y, WANG Y, CHEN T, et al. Deep functional analysis of synII, a 770-kilobase synthetic yeast chromosome[J]. Science, 2017, 355 (6329): eaaf4791.
57	WU Y, LI B Z, ZHAO M, et al. Bug mapping and fitness testing of chemically synthesized chromosome X[J]. Science, 2017, 355 (6329): eaaf4706.
58	ZHANG W, ZHAO G, LUO Z, et al. Engineering the ribosomal DNA in a megabase synthetic chromosome[J]. Science, 2017, 355 (6329): eaaf3981.
59	XIE Z X, LIU D, LI B Z, et al. Design and chemical synthesis of eukaryotic chromosomes[J]. Chemical Society Reviews, 2017, 46 (23): 7191-7207.
60	ISAACS F J, CARR P A, WANG H H, et al. Precise manipulation of chromosomes in vivo enables genome-wide codon replacement[J]. Science, 2011, 333 (6040): 348-353.
61	KOMOR A C, KIM Y B, PACKER M S, et al. Programmable editing of a target base in genomic DNA without double-stranded DNA cleavage[J]. Nature, 2016, 533 (7603): 420-424.
62	GAUDELLI N M, KOMOR A C, REES H A, et al. Programmable base editing of A•T to G•C in genomic DNA without DNA cleavage[J]. Nature, 2017, 551 (7681): 464-471.
63	BAO Z, HAMEDIRAD M, XUE P, et al. Genome-scale engineering of Saccharomyces cerevisiae with single-nucleotide precision[J]. Nature Biotechnology, 2018, 36 (6): 505-508.
64	JIA B, WU Y, LI B Z, et al. Precise control of SCRaMbLE in synthetic haploid and diploid yeast[J]. Nature Communications, 2018, 9 (1): 1933.
65	WANG J, XIE Z X, MA Y, et al. Ring synthetic chromosome ⅤSCRaMbLE[J]. Nature Communications, 2018, 9 (1): 3783.
66	NAKAMURA C E, WHITED G M. Metabolic engineering for the microbial production of 1,3-propanediol[J]. Current Opinion in Biotechnology, 2003, 14 (5): 454-459.
67	RO D K, PARADISE E M, OUELLET M, et al. Production of the antimalarial drug precursor artemisinic acid in engineered yeast[J]. Nature, 2006, 440 (7086): 940-943.
68	MARTIN V J, PITERA D J, WITHERS S T, et al. Engineering a mevalonate pathway in Escherichia coli for production of terpenoids[J]. Nature Biotechnology, 2003, 21 (7): 796-802.
69	NAKAGAWA A, MATSUMURA E, KOYANAGI T, et al. Total biosynthesis of opiates by stepwise fermentation using engineered Escherichia coli[J]. Nature Communications, 2016, 7: 10390.
70	LUO X, REITER M A, D'ESPAUX L, et al. Complete biosynthesis of cannabinoids and their unnatural analogues in yeast[J]. Nature, 2019, 567 (7746): 123-126.
71	CHEN X L, GAO C, GUO L, et al. DCEO biotechnology: tools to design, construct, evaluate, and optimize the metabolic pathway for biosynthesis of chemicals[J]. Chemical Reviews, 2018, 118 (1): 4-72.
72	DU Y L, ALKHALAF L M, RYAN K S. In vitro reconstitution of indolmycin biosynthesis reveals the molecular basis of oxazolinone assembly[J]. PNAS, 2015, 112 (9): 2717-2722.
73	LUO Y, HUANG H, LIANG J, et al. Activation and characterization of a cryptic polycyclic tetramate macrolactam biosynthetic gene cluster[J]. Nature Communications, 2013, 4: 2894.
74	TU L, SU P, ZHANG Z, et al. GenomGenome of Tripterygium wilfordii and identification of cytochrome P450 involved in triptolide biosynthesis [J]. Nature Communications, 2020, 11 (1): 971.
75	LAU W, SATTELY E S. Six enzymes from mayapple that complete the biosynthetic pathway to the etoposide aglycone[J]. Science, 2015, 349 (6253): 1224-1228.
76	HAMMER S C, KNIGHT A M, ARNOLD F H. Design and evolution of enzymes for non-natural chemistry[J]. Current Opinion in Green and Sustainable Chemistry, 2017, 7: 23-30.
77	CHEN K, ARNOLD F H. Engineering new catalytic activities in enzymes[J]. Nature Catalysis, 2020, 3 (3): 203-213.
78	LEAVER-FAY A, TYKA M, LEWIS S M, et al. ROSETTA3: an object-oriented software suite for the simulation and design of macromolecules[J]. Methods Enzymol, 2011, 487: 545-574.
79	DOU J Y, VOROBIEVA A A, SHEFFLER W, et al. De novo design of a fluorescence-activating beta-barrel[J]. Nature, 2018, 561 (7724): 485-491.
80	BOYKEN S E, BENHAIM M A, BUSCH F, et al. De novo design of tunable, pH-driven conformational changes[J]. Science, 2019, 364 (6441): 658-664.
81	CHEN Z B, BOYKEN S E, JIA M X, et al. Programmable design of orthogonal protein heterodimers[J]. Nature, 2019, 565 (7737): 106-111.
82	HAMMER S C, KUBIK G, WATKINS E, et al. Anti-Markovnikov alkene oxidation by metal-oxo-mediated enzyme catalysis[J]. Science, 2017, 358 (6360): 215-218.
83	KAN S B J, HUANG X, GUMULYA Y, et al. Genetically programmed chiral organoborane synthesis[J]. Nature, 2017, 552 (7683): 132-136.
84	KAN S B, LEWIS R D, CHEN K, et al. Directed evolution of cytochrome c for carbon-silicon bond formation: bringing silicon to life[J]. Science, 2016, 354 (6315): 1048-1051.
85	IGNEA C, PONTINI M, MOTAWIA M S, et al. Synthesis of 11-carbon terpenoids in yeast using protein and metabolic engineering[J]. Nature Chemical Biology, 2018, 14 (12): 1090-1098.
86	FURUBAYASHI M, IKEZUMI M, TAKAICHI S, et al. A highly selective biosynthetic pathway to non-natural C50 carotenoids assembled from moderately selective enzymes[J]. Nature Communications, 2015, 6: 7534.
87	XIONG M, SCHNEIDERMAN D K, BATES F S, et al. Scalable production of mechanically tunable block polymers from sugar[J]. PNAS, 2014, 111 (23): 8357-8362.
88	TAI Y S, XIONG M, JAMBUNATHAN P, et al. Engineering nonphosphorylative metabolism to generate lignocellulose-derived products[J]. Nature Chemical Biology, 2016, 12 (4): 247-253.
89	YAO Y F, WANG C S, QIAO J, et al. Metabolic engineering of Escherichia coli for production of salvianic acid A via an artificial biosynthetic pathway[J]. Metabolic Engineering, 2013, 19: 79-87.
90	AJIKUMAR P K, XIAO W H, TYO K E, et al. Isoprenoid pathway optimization for Taxol precursor overproduction in Escherichia coli[J]. Science, 2010, 330 (6000): 70-74.
91	XU P, GU Q, WANG W, et al. Modular optimization of multi-gene pathways for fatty acids production in E. coli[J]. Nature Communications, 2013, 4: 1409.
92	QIN J, ZHOU Y J, KRIVORUCHKO A, et al. Modular pathway rewiring of Saccharomyces cerevisiae enables high-level production of L-ornithine[J]. Nature Communications, 2015, 6: 8224.
93	XU P, QIAO K, AHN W S, et al. Engineering Yarrowia lipolytica as a platform for synthesis of drop-in transportation fuels and oleochemicals[J]. PNAS, 2016, 113 (39): 10848-10853.
94	FANG H, LI D, KANG J, et al. Metabolic engineering of Escherichia coli for de novo biosynthesis of vitamin B₁₂[J]. Nature Communications, 2018, 9 (1): 4917.
95	SRINIVASAN P, SMOLKE C D. Engineering a microbial biosynthesis platform for de novo production of tropane alkaloids[J]. Nature Communications, 2019, 10 (1): 3634.
96	SWEETLOVE L J, FERNIE A R. The role of dynamic enzyme assemblies and substrate channelling in metabolic regulation[J]. Nature Communications, 2018, 9 (1): 2136.
97	HAMMER S K, AVALOS J L. Harnessing yeast organelles for metabolic engineering[J]. Nature Chemical Biology, 2017, 13 (8): 823-832.
98	BERNER N, REUTTER K R, WOLF D H. Protein quality control of the endoplasmic reticulum and ubiquitin-proteasome-triggered degradation of aberrant proteins: yeast pioneers the path[J]. Annual Review of Biochemistry, 2018, 87: 751-782.
99	CHEN S J, WU X, WADAS B, et al. An N-end rule pathway that recognizes proline and destroys gluconeogenic enzymes[J]. Science, 2017, 355 (6323): aal3655.
100	PENG B, NIELSEN L K, KAMPRANIS S C, et al. Engineered protein degradation of farnesyl pyrophosphate synthase is an effective regulatory mechanism to increase monoterpene production in Saccharomyces cerevisiae[J]. Metabolic Engineering, 2018, 47: 83-93.
101	PENG B, PLAN M R, CHRYSANTHOPOULOS P, et al. A squalene synthase protein degradation method for improved sesquiterpene production in Saccharomyces cerevisiae[J]. Metabolic Engineering, 2017, 39: 209-219.
102	MORSUT L, ROYBAL K T, XIONG X, et al. Engineering customized cell sensing and response behaviors using synthetic notch receptors[J]. Cell, 2016, 164 (4): 780-791.
103	KENT R, DIXON N. Systematic evaluation of genetic and environmental factors affecting performance of translational riboswitches[J]. ACS Synthetic Biology, 2019, 8 (4): 884-901.
104	DAHLMAN J E, ABUDAYYEH O O, JOUNG J, et al. Orthogonal gene knockout and activation with a catalytically active Cas9 nuclease[J]. Nature Biotechnology, 2015, 33 (11): 1159-1161.
105	PANDIT A V, SRINIVASAN S, MAHADEVAN R. Redesigning metabolism based on orthogonality principles[J]. Nature Communications, 2017, 8: 15188.
106	DE LORENZO V. Beware of metaphors: chasses and orthogonality in synthetic biology[J]. Bioengineered Bugs, 2011, 2 (1): 3-7.
107	LIU H, LU T. Autonomous production of 1,4-butanediol via a de novo biosynthesis pathway in engineered Escherichia coli[J]. Metabolic Engineering, 2015, 29: 135-141.
108	IGNEA C, RAADAM M H, MOTAWIA M S, et al. Orthogonal monoterpenoid biosynthesis in yeast constructed on an isomeric substrate[J]. Nature Communications, 2019, 10 (1): 3799.
109	JI D, WANG L, HOU S, et al. Creation of bioorthogonal redox systems depending on nicotinamide flucytosine dinucleotide[J]. Journal of the American Chemical Society, 2011, 133 (51): 20857-20862.
110	LIU Y, FENG Y, WANG L, et al. Structural insights into phosphite dehydrogenase variants favoring a non-natural redox cofactor[J]. ACS Catalysis, 2019, 9 (3): 1883-1887.
111	WANG L, JI D, LIU Y, et al. Synthetic cofactor-linked metabolic circuits for selective energy transfer[J]. ACS Catalysis, 2017, 7 (3): 1977-1983.
112	GUO X, LIU Y, WANG Q, et al. Non-natural cofactor and formate-driven reductive carboxylation of pyruvate[J]. Angewandte Chemie International Edition, 2020, 59 (8): 3143-3146.
113	LIU G S, LI T, ZHOU W, et al. The yeast peroxisome: a dynamic storage depot and subcellular factory for squalene overproduction[J]. Metabolic Engineering, 2020, 57: 151-161.
114	BALDI N, DYKSTRA J C, LUTTIK M A H, et al. Functional expression of a bacterial alpha-ketoglutarate dehydrogenase in the cytosol of Saccharomyces cerevisiae[J]. Metabolic Engineering, 2019, 56: 190-197.
115	LV X, WANG F, ZHOU P, et al. Dual regulation of cytoplasmic and mitochondrial acetyl-CoA utilization for improved isoprene production in Saccharomyces cerevisiae[J]. Nature Communications, 2016, 7: 12851.
116	SADRE R, KUO P, CHEN J, et al. Cytosolic lipid droplets as engineered organelles for production and accumulation of terpenoid biomaterials in leaves[J]. Nature Communications, 2019, 10 (1): 853.
117	ZHAO C, KIM Y, ZENG Y, et al. Co-compartmentation of terpene biosynthesis and storage via synthetic droplet[J]. ACS Synthetic Biology, 2018, 7 (3): 774-781.
118	ZHOU Y J, BUIJS N A, ZHU Z, et al. Harnessing yeast peroxisomes for biosynthesis of fatty-acid-derived biofuels and chemicals with relieved side-pathway competition[J]. Journal of the American Chemical Society, 2016, 138 (47): 15368-15377.
119	KIM J E, JANG I S, SON S H, et al. Tailoring the Saccharomyces cerevisiae endoplasmic reticulum for functional assembly of terpene synthesis pathway[J]. Metabolic Engineering, 2019, 56: 50-59.
120	CHEN R, YANG S, ZHANG L, et al. Advanced strategies for production of natural products in yeast[J]. iScience, 2020, 23 (3): 100879.
121	KLEI I J VAN DER, VEENHUIS M. Yeast peroxisomes: function and biogenesis of a versatile cell organelle[J]. Trends Microbiol., 1997, 5 (12): 502-509.
122	CROSS L L, PAUDYAL R, KAMISUGI Y, et al. Towards designer organelles by subverting the peroxisomal import pathway[J]. Nature Communications, 2017, 8 (1): 454.
123	STEINKUHLER J, KNORR R L, ZHAO Z, et al. Controlled division of cell-sized vesicles by low densities of membrane-bound proteins[J]. Nature Communications, 2020, 11 (1): 905.
124	KERFELD C A, AUSSIGNARGUES C, ZARZYCKI J, et al. Bacterial microcompartments[J]. Nat. Rev. Microbiol., 2018, 16 (5): 277-290.
125	HAGEN A, SUTTER M, SLOAN N, et al. Programmed loading and rapid purification of engineered bacterial microcompartment shells[J]. Nature Communications, 2018, 9 (1): 2881.
126	LEE M J, MANTELL J, BROWN I R, et al. De novo targeting to the cytoplasmic and luminal side of bacterial microcompartments[J]. Nature Communications, 2018, 9 (1): 3413.
127	FERLEZ B, SUTTER M, KERFELD C A. A designed bacterial microcompartment shell with tunable composition and precision cargo loading[J]. Metabolic Engineering, 2019, 54: 286-291.
128	GIESSEN T W, SILVER P A. Widespread distribution of encapsulin nanocompartments reveals functional diversity[J]. Nature Microbiology, 2017, 2 (6): 17029.
129	BUGAJ L J, CHOKSI A T, MESUDA C K, et al. Optogenetic protein clustering and signaling activation in mammalian cells[J]. Nature Methods, 2013, 10 (3): 249-252.
130	GIL A A, LAPTENOK S P, IULIANO J N, et al. Photoactivation of the BLUF protein PixD Probed by the site-specific incorporation of fluorotyrosine residues[J]. Journal of the American Chemical Society, 2017, 139 (41): 14638-14648.
131	LAU Y H, GIESSEN T W, ALTENBURG W J, et al. Prokaryotic nanocompartments form synthetic organelles in a eukaryote[J]. Nature Communications, 2018, 9 (1): 1311.
132	ZHAO E M, SUEK N, WILSON M Z, et al. Light-based control of metabolic flux through assembly of synthetic organelles[J]. Nature Chemical Biology, 2019, 15 (6): 589-597.
133	RUGBJERG P, SOMMER M O A. Overcoming genetic heterogeneity in industrial fermentations[J]. Nature Biotechnology, 2019, 37 (8): 869-876.
134	RUGBJERG P, SARUP-LYTZEN K, NAGY M, et al. Synthetic addiction extends the productive life time of engineered Escherichia coli populations[J]. PNAS, 2018, 115 (10): 2347-2352.
135	XIAO Y, BOWEN C H, LIU D, et al. Exploiting nongenetic cell-to-cell variation for enhanced biosynthesis[J]. Nature Chemical Biology, 2016, 12 (5): 339-344.
136	RAMAN S, ROGERS J K, TAYLOR N D, et al. Evolution-guided optimization of biosynthetic pathways[J]. PNAS, 2014, 111 (50): 17803-17808.
137	GUPTA A, REIZMAN I M, REISCH C R, et al. Dynamic regulation of metabolic flux in engineered bacteria using a pathway-independent quorum-sensing circuit[J]. Nature Biotechnology, 2017, 35 (3): 273-279.
138	DAHL R H, ZHANG F, ALONSO-GUTIERREZ J, et al. Engineering dynamic pathway regulation using stress-response promoters[J]. Nature Biotechnology, 2013, 31 (11): 1039-1046.
139	LIANG C, ZHANG X, WU J, et al. Dynamic control of toxic natural product biosynthesis by an artificial regulatory circuit[J]. Metabolic Engineering, 2020, 57: 239-246.
140	CERONI F, BOO A, FURINI S, et al. Burden-driven feedback control of gene expression[J]. Nature Methods, 2018, 15 (5): 387-393.
141	JULLESSON D, DAVID F, PFLEGER B, et al. Impact of synthetic biology and metabolic engineering on industrial production of fine chemicals[J]. Biotechnology Advances, 2015, 33 (7): 1395-1402.
142	SANDBERG T E, SALAZAR M J, WENG L L, et al. The emergence of adaptive laboratory evolution as an efficient tool for biological discovery and industrial biotechnology[J]. Metabolic Engineering, 2019, 56: 1-16.
143	FLETCHER E, FEIZI A, BISSCHOPS M M M, et al. Evolutionary engineering reveals divergent paths when yeast is adapted to different acidic environments[J]. Metabolic Engineering, 2017, 39: 19-28.
144	DEATHERAGE D E, KEPNER J L, BENNETT A F, et al. Specificity of genome evolution in experimental populations of Escherichia coli evolved at different temperatures[J]. PNAS, 2017, 114 (10): E1904-E1912.
145	MUNDHADA H, SEOANE J M, SCHNEIDER K, et al. Increased production of L-serine in Escherichia coli through adaptive laboratory evolution[J]. Metabolic Engineering, 2017, 39: 141-150.
146	RODRíGUEZ-VERDUGO A, CARRILLO-CISNEROS D, GONZáLEZ-GONZáLEZ A, et al. Different tradeoffs result from alternate genetic adaptations to a common environment[J]. PNAS, 2014, 111 (33): 12121.
147	CASPETA L, CHEN Y, GHIACI P, et al. Altered sterol composition renders yeast thermotolerant[J]. Science, 2014, 346 (6205): 75-78.
148	THORWALL S, SCHWARTZ C, CHARTRON J W, et al. Stress-tolerant non-conventional microbes enable next-generation chemical biosynthesis[J]. Nature Chemical Biology, 2020, 16 (2): 113-121.
149	CALERO P, NIKEL P I. Chasing bacterial chassis for metabolic engineering: a perspective review from classical to non-traditional microorganisms[J]. Microbial Biotechnology, 2019, 12 (1): 98-124.
150	KUMAR V, PARK S. Potential and limitations of Klebsiella pneumoniae as a microbial cell factory utilizing glycerol as the carbon source[J]. Biotechnology Advances, 2018, 36 (1): 150-167.
151	CHARUBIN K, BENNETT R K, FAST A G, et al. Engineering Clostridium organisms as microbial cell-factories: challenges & opportunities[J]. Metabolic Engineering, 2018, 50: 173-191.
152	TAN D, WU Q, CHEN J C, et al. Engineering Halomonas TD01 for the low-cost production of polyhydroxyalkanoates[J]. Metabolic Engineering, 2014, 26: 34-47.
153	BRENNER K, YOU L, ARNOLD F H. Engineering microbial consortia: a new frontier in synthetic biology[J]. Trends in Biotechnology, 2008, 26 (9): 483-489.
154	JONES J A, WANG X. Use of bacterial co-cultures for the efficient production of chemicals[J]. Current Opinion in Biotechnology, 2018, 53: 33-38.
155	SONG H, DING M Z, JIA X Q, et al. Synthetic microbial consortia: from systematic analysis to construction and applications[J]. Chemical Society Reviews, 2014, 43 (20): 6954-6981.
156	HAYS S G, PATRICK W G, ZIESACK M, et al. Better together: engineering and application of microbial symbioses[J]. Current Opinion in Biotechnology, 2015, 36: 40-49.
157	KONG W T, MELDGIN D R, COLLINS J J, et al. Designing microbial consortia with defined social interactions[J]. Nature Chemical Biology, 2018, 14 (8): 821-829.
158	JAROSZ D F, BROWN J C S, WALKER G A, et al. Cross-kingdom chemical communication drives a heritable, mutually beneficial prion-based transformation of metabolism[J]. Cell, 2014, 158 (5): 1083-1093.
159	ZELEZNIAK A, ANDREJEV S, PONOMAROVA O, et al. Metabolic dependencies drive species co-occurrence in diverse microbial communities[J]. PNAS, 2015, 112 (20): 6449-6454.
160	WINTERMUTE E H, SILVER P A. Dynamics in the mixed microbial concourse[J]. Genes & Development, 2010, 24 (23): 2603-2614.
161	BRENNER K, KARIG D K, WEISS R, et al. Engineered bidirectional communication mediates a consensus in a microbial biofilm consortium[J]. PNAS, 2007, 104 (44): 17300-17304.
162	FAUST K, RAES J. Microbial interactions: from networks to models[J]. Nature Reviews Microbiology, 2012, 10 (8): 538-550.
163	MCCARTY N S, LEDESMA-AMARO R. Synthetic biology tools to engineer microbial communities for biotechnology[J]. Trends in Biotechnology, 2019, 37 (2): 181-197.
164	SCOTT S R, DIN M O, BITTIHN P, et al. A stabilized microbial ecosystem of self-limiting bacteria using synthetic quorum-regulated[J]. Nature Microbiology, 2017, 2 (8): 17083.
165	SHOU W Y, RAM S, VILAR J M G. Synthetic cooperation in engineered yeast populations[J]. PNAS, 2007, 104 (6): 1877-1882.
166	HU B, DU J, ZOU R Y, et al. An environment-sensitive synthetic microbial ecosystem[J]. PLoS One, 2010, 5 (5): e10619.
167	WANG E X, DING M Z, MA Q, et al. Reorganization of a synthetic microbial consortium for one-step vitamin C fermentation[J]. Microbial Cell Factories, 2016, 15: 21.
168	ROELL G W, ZHA J, CARR R R, et al. Engineering microbial consortia by division of labor[J]. Microbial Cell Factories, 2019, 18 (1): 35.
169	LIU Y, DING M Z, LING W, et al. A three-species microbial consortium for power generation[J]. Energy & Environmental Science, 2017, 10 (7): 1600-1609.
170	ZHOU K, QIAO K J, EDGAR S, et al. Distributing a metabolic pathway among a microbial consortium enhances production of natural products[J]. Nature Biotechnology, 2015, 33 (4): 377-383.
171	JONES J A, VERNACCHIO V R, SINKOE A L, et al. Experimental and computational optimization of an Escherichia coli co-culture for the efficient production of flavonoids[J]. Metabolic Engineering, 2016, 35: 55-63.
172	PANDE S, SHITUT S, FREUND L, et al. Metabolic cross-feeding via intercellular nanotubes among bacteria[J]. Nature Communications, 2015, 6: 6238.
173	BENOMAR S, RANAVA D, CARDENAS M L, et al. Nutritional stress induces exchange of cell material and energetic coupling between bacterial species[J]. Nature Communications, 2015, 6: 6283.
174	WEBER W, BABA M, FUSSENEGGER M. Synthetic ecosystems based on airborne inter- and intrakingdom communication[J]. PNAS, 2007, 104 (25): 10435-10440.
175	CHEN Z Y, SUN X X, LI Y, et al. Metabolic engineering of Escherichia coli for microbial synthesis of monolignols[J]. Metabolic Engineering, 2017, 39: 102-109.
176	JONES J A, VERNACCHIO V R, COLLINS S M, et al. Complete biosynthesis of Anthocyanins using E. coli polycultures[J]. Mbio., 2017, 8 (3): e00621-00617.
177	QIAN X, CHEN L, SUI Y, et al. Biotechnological potential and applications of microbial consortia[J]. Biotechnology Advances, 2020, 40: 107500.
178	SGOBBA E, WENDISCH V F. Synthetic microbial consortia for small molecule production[J]. Current Opinion in Biotechnology, 2019, 62: 72-79.
179	ZHANG H, PEREIRA B, LI Z, et al. Engineering Escherichia coli coculture systems for the production of biochemical products[J]. PNAS, 2015, 112 (27): 8266-8271.
180	LIU X, LI X B, JIANG J L, et al. Convergent engineering of syntrophic Escherichia coli coculture for efficient production of glycosides[J]. Metabolic Engineering, 2018, 47: 243-253.
181	ZHAO C H, SINUMVAYO J P, ZHANG Y P, et al. Design and development of a "Y-shaped" microbial consortium capable of simultaneously utilizing biomass sugars for efficient production of butanol[J]. Metabolic Engineering, 2019, 55: 111-119.
182	AKDEMIR H, SILVA A, ZHA J, et al. Production of pyranoanthocyanins using Escherichia coli co-cultures[J]. Metabolic Engineering, 2019, 55: 290-298.
183	GUO X Y, LI Z H, WANG X N, et al. De novo phenol bioproduction from glucose using biosensor-assisted microbial coculture engineering[J]. Biotechnology and Bioengineering, 2019, 116 (12): 3349-3359.
184	MINTY J J, SINGER M E, SCHOLZ S A, et al. Design and characterization of synthetic fungal-bacterial consortia for direct production of isobutanol from cellulosic biomass[J]. PNAS, 2013, 110 (36): 14592-14597.
185	SCHOLZ S A, GRAVES I, MINTY J J, et al. Production of cellulosic organic acids via synthetic fungal consortia[J]. Biotechnology and Bioengineering, 2018, 115 (4): 1096-1100.
186	TSAI S L, GOYAL G, CHEN W. Surface display of a functional minicellulosome by intracellular complementation using a synthetic yeast consortium and its application to cellulose hydrolysis and ethanol production[J]. Applied and Environmental Microbiology, 2010, 76 (22): 7514-7520.
187	WEN Z Q, MINTON N P, ZHANG Y, et al. Enhanced solvent production by metabolic engineering of a twin-clostridial consortium[J]. Metabolic Engineering, 2017, 39: 38-48.
188	GILBERT E S, WALKER A W, KEASLING J D. A constructed microbial consortium for biodegradation of the organophosphorus insecticide parathion[J]. Applied Microbiology and Biotechnology, 2003, 61 (1): 77-81.
189	ZHANG H, YANG C, LI C K, et al. Functional assembly of a microbial consortium with autofluorescent and mineralizing activity for the biodegradation of organophosphates[J]. Journal of Agricultural and Food Chemistry, 2008, 56 (17): 7897-7902.
190	MARTINEZ I, MOHAMED M E S, ROZAS D, et al. Engineering synthetic bacterial consortia for enhanced desulfurization and revalorization of oil sulfur compounds[J]. Metabolic Engineering, 2016, 35: 46-54.
191	IBRAR M, ZHANG H. Construction of a hydrocarbon-degrading consortium and characterization of two new lipopeptides biosurfactants[J]. Science of the Total Environment, 2020, 714: 136400.
192	ERLICH Y, ZIELINSKI D. DNA Fountain enables a robust and efficient storage architecture[J]. Science, 2017, 355 (6328): 950-953.
193	ORGANICK L, ANG S D, CHEN Y J, et al. Random access in large-scale DNA data storage[J]. Nature Biotechnology, 2018, 36 (7): 660-660.
194	CEZE L, NIVALA J, STRAUSS K. Molecular digital data storage using DNA[J]. Nature Reviews Genetics, 2019, 20 (8): 456-466.
195	GOLDMAN N, BERTONE P, CHEN S Y, et al. Towards practical, high-capacity, low-maintenance information storage in synthesized DNA[J]. Nature, 2013, 494 (7435): 77-80.
196	CHURCH G M, GAO Y, KOSURI S. Next-generation digital information storage in DNA[J]. Science, 2012, 337 (6102): 1628.
197	KOCH J, GANTENBEIN S, MASANIA K, et al. A DNA-of-things storage architecture to create materials with embedded memory[J]. Nature Biotechnology, 2020, 38 (1): 39-43.
198	GRASS R N, HECKEL R, PUDDU M, et al. Robust chemical preservation of digital information on DNA in silica with error-correcting codes[J]. Angewandte Chemie-International Edition, 2015, 54 (8): 2552-2555.
199	TAKAHASHI C N, NGUYEN B H, STRAUSS K, et al. Demonstration of end-to-end automation of DNA data storage[J]. Scientific Reports, 2019, 9 (1): 4998.
200	CHOI Y, RYU T, LEE A C, et al. High information capacity DNA-based data storage with augmented encoding characters using degenerate bases[J]. Scientific Reports, 2019, 9 (1): 6582.
201	ANAVY L, VAKNIN I, ATAR O, et al. Data storage in DNA with fewer synthesis cycles using composite DNA letters[J]. Nature Biotechnology, 2019, 37 (10): 1237.
202	陈为刚, 黄刚, 李炳志, 等. 音视频文件的DNA信息存储[J]. 中国科学:生命科学, 2020, 50 (1): 81-85.
	CHEN W G, HUANG G, LI B Z, et al. DNA information storage for audio and video files (in Chinese)[J]. SCIENTIA SINICA Vitae, 2020, 50(1): 81-85.
203	ROTHEMUND P W K. Folding DNA to create nanoscale shapes and patterns[J]. Nature, 2006, 440 (7082): 297-302.
204	钱璐璐, 汪颖, 张钊, 等. DNA纳米结构仿中国地图[J]. 科学通报， 2006, 51 (24): 2860-2863.
	QIAN L L, WANG Y, ZHANG Z, et al. Imating China map with DNA nanostructure[J]. Chinese Science Bulletin， 2006， 51 (24): 2860-2863.
205	DOUGLAS S M, CHOU J J, SHIH W M. DNA-nanotube-induced alignment of membrane proteins for NMR structure determination[J]. PNAS, 2007, 104 (16): 6644-6648.
206	ANDERSEN E S, DONG M, NIELSEN M M, et al. Self-assembly of a nanoscale DNA box with a controllable lid[J]. Nature, 2009, 459 (7243): 73-76.
207	HAN D R, PAL S, NANGREAVE J, et al. DNA origami with complex curvatures in three-dimensional space[J]. Science, 2011, 332 (6027): 342-346.
208	DU Y, JIANG Q, BEZIERE N, et al. DNA-nanostructure-gold-nanorod hybrids for enhanced in vivo optoacoustic imaging and photothermal therapy[J]. Advanted Materials, 2016, 28 (45): 10000-10007.
209	ZHANG Y N, WANG F, CHAO J, et al. DNA origami cryptography for secure communication[J]. Nature Communications, 2019, 10 (1): 5469.
210	BAZRAFSHAN A, MEYER T A, SU H Q, et al. Tunable DNA origami motors translocate ballistically over μm distances at nm/s speeds[J]. Angewandte Chemie-International Edition, 2020, 132: 2-10.
211	YOUNG T S, SCHULTZ P G. Beyond the canonical 20 amino acids: expanding the genetic lexicon[J]. Journal of Biological Chemistry, 2010, 285 (15): 11039-11044.
212	DEEPANKUMAR K, SHON M, NADARAJAN S P, et al. Enhancing thermostability and organic solvent tolerance of w-transaminase through global incorporation of fluorotyrosine[J]. Advanced Synthesis & Catalysis, 2014, 356 (5): 993-998.
213	DRIENOVSKA I, MAYER C, DULSON C, et al. A designer enzyme for hydrazone and oxime formation featuring an unnatural catalytic aniline residue[J]. Nature Chemistry, 2018, 10 (9): 946-952.
214	ROVNER A J, HAIMOVICH A D, KATZ S R, et al. Recoded organisms engineered to depend on synthetic amino acids[J]. Nature, 2015, 518 (7537): 89-93.
215	MANDELL D J, LAJOIE M J, MEE M T, et al. Biocontainment of genetically modified organisms by synthetic protein design[J]. Nature, 2015, 518 (7537): 55-60.
216	YU Y, LV X, LI J, et al. Defining the role of tyrosine and rational tuning of oxidase activity by genetic incorporation of unnatural tyrosine analogs[J]. Journal of the American Chemical Society, 2015, 137 (14): 4594-4597.
217	YOUNG D D, SCHULTZ P G. Playing with the molecules of life[J]. ACS Chemical Biology, 2018, 13 (4): 854-870.
218	ZHANG Y, PTACIN J L, FISCHER E C, et al. A semi-synthetic organism that stores and retrieves increased genetic information[J]. Nature, 2017, 551 (7682): 644-647.
219	DUNN M R, JIMENEZ R M, CHAPUT J C. Analysis of aptamer discovery and technology[J]. Nature Reviews Chemistry, 2017, 1 (10): 0076.
220	CHEN T, HONGDILOKKUL N, LIU Z X, et al. The expanding world of DNA and RNA[J]. Current Opinion in Chemical Biology, 2016, 34: 80-87.
221	HOSHIKA S, LEAL N A, KIM M J, et al. Hachimoji DNA and RNA: a genetic system with eight building blocks[J]. Science, 2019, 363 (6429): 884-887.
222	EREMEEVA E, HERDEWIJN P. Non canonical genetic material[J]. Current Opinion in Biotechnology, 2019, 57: 25-33.
223	KUK S K, SINGH R K, NAM D H, et al. Photoelectrochemical reduction of carbon dioxide to methanol through a highly efficient enzyme cascade[J]. Angewandte Chemie-International Edition, 2017, 56 (14): 3827-3832.
224	SOKOL K P, ROBINSON W E, WARNAN J, et al. Bias-free photoelectrochemical water splitting with photosystem II on a dye-sensitized photoanode wired to hydrogenase[J]. Nature Energy, 2018, 3 (11): 944-951.
225	SAKIMOTO K K, WONG A B, YANG P D. Self-photosensitization of nonphotosynthetic bacteria for solar-to-chemical production[J]. Science, 2016, 351 (6268): 74-77.
226	GUO J L, SUASTEGUI M, SAKIMOTO K K, et al. Light-driven fine chemical production in yeast biohybrids[J]. Science, 2018, 362 (6416): 813-816.
227	WANG X Y, PU J H, AN B L, et al. Programming cells for dynamic assembly of Iiorganic nano-objects with spatiotemporal control[J]. Advanced Materials, 2018, 30 (16): e1705968.
228	WANG X Y, PU J H, LIU Y, et al. Immobilization of functional nano-objects in living engineered bacterial biofilms for catalytic applications[J]. National Science Review, 2019, 6 (5): 929-943.
229	JIN X, HONG S H. Cell-free protein synthesis for producing ‘difficult-to-express’ proteins[J]. Biochemical Engineering Journal, 2018, 138: 156-164.
230	SHINODA T, SHINYA N, ITO K, et al. Cell-free methods to produce structurally intact mammalian membrane proteins[J]. Scientific Reports, 2016, 6: 30442.
231	MARSHALL R, MAXWELL C S, COLLINS S P, et al. Rapid and scalable characterization of CRISPR technologies using an E. coli cell-free transcription-translation system[J]. Molecular Cell, 2018, 69 (1): 146-157.e3.
232	MARTIN R W, MAJEWSKA N I, CHEN C X, et al. Development of a CHO-based cell-free platform for synthesis of active monoclonal antibodies[J]. ACS Synthetic Biology, 2017, 6 (7): 1370-1379.
233	RAMOS-BENITEZ M J, LOPEZ-CRUZ L M, AGUAYO V, et al. Cell-free expression, purification and immunoreactivity assessment of recombinant Fasciola hepatica saposin-like protein-2[J]. Molecular Biology Reports, 2018, 45 (5): 1551-1556.
234	GAO W, CHO E, LIU Y, et al. Advances and challenges in cell-free incorporation of unnatural amino acids into proteins[J]. Front Pharmacol, 2019, 10: 611.
235	ZIMMERMAN E S, HEIBECK T H, GILL A, et al. Production of site-specific antibody-drug conjugates using optimized non-natural amino acids in a cell-free expression system[J]. Bioconjugate Chemistry, 2014, 25 (2): 351-361.
236	HOFFMANN B, LOHR F, LAGUERRE A, et al. Protein labeling strategies for liquid-state NMR spectroscopy using cell-free synthesis[J]. Progress in Nuclear Magnetic Resonance Spectroscopy, 2018, 105: 1-22.
237	PEREZ J G, STARK J C, JEWETT M C. Cell-free synthetic biology: engineering beyond the cell[J]. Cold Spring Harbor Perspectives in Biology, 2016, 8 (12): a023853.
238	ADACHI J, KATSURA K, SEKI E, et al. Cell-free protein synthesis using S30 extracts from Escherichia coli RFzero strains for efficient incorporation of non-natural amino acids into proteins[J]. International Journal of Molecular Sciences, 2019, 20 (3): 492.

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