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Table of Content
25 January 2017, Volume 33 Issue 1
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Orignal Article
Metabolite Biosensor:A Useful Synthetic Biology Tool to Assist the Construction of Microbial Cell Factory
ZHOU Yi-kang, WU Yi-nan, WANG Tian-min, ZHENG Xiang, XING Xin-hui, ZHANG Chong
2017, 33(1): 1-11. doi:
10.13560/j.cnki.biotech.bull.1985.2017.01.001
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Metabolite biosensor,which represents a useful synthetic biology tool,can sense the concentration of intracellular metabolites and then transfer it into specific signal output,showing great potential in the construction of microbial cell factory. It generally consists of two components,biological recognition element and signal output element. The former,typified by regulatory elements,is ubiquitous in nature,such as transcription factor and riboswitch,and has diverse response mechanism. The output can be fluorescence signals,fitness,pathway activation or silence,and so on. Here we reviewed recent progresses about applications of metabolite biosensor in the construction of microbial cell factory,including high-throughput screen/selection,dynamic control and non-genetic selection. We also discussed the effects of the metabolite biosensor performance in application and focused on the opportunities and challenges we might meet in practice.
Key Technologies and Applications of Synthetic Biology
YANG Ju, DENG Yu
2017, 33(1): 12-23. doi:
10.13560/j.cnki.biotech.bull.1985.2017.01.002
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Synthetic biology is to reconstruct the existing natural system by building standardized components and modules and synthesize new artificial life system from scratch under the guidance of engineering design ideas.With the help of gene recombination technology and genome editing technology,we can program life system to comply special functions. After realizing modular analysis of metabolic pathways and optimizing the combination between the components we can synthesize chemicals with excellent patterns. At present,people have made great progress in the fields of energy,chemical industry and medicine. Synthetic biology will bring great changes to people’s life and will continue to be the hotspot of research.The review describes the major applications and the key technologies of synthetic biology in genome editing and modular representation.
Research Advance on Biosynthesis of Aromatic Amino Acids and Their Derivatives
SHEN Xiao-lin, YUAN Qi-peng
2017, 33(1): 24-34. doi:
10.13560/j.cnki.biotech.bull.1985.2017.01.003
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Bioengineering has been used to produce various of value-added products including fine chemicals,bulk chemicals,pharmaceuticals and nutraceuticals using sustainable materials. Synthetic biology is the foundation of bioengineering which combined functional genomics,computational biology with system biology to createnovel products and optimize bioprocess. In the past decades,many powerful tools developed from synthetic biology has been used in microorganisms,plants and animals. In this review,we summarize the synthesis of aromatic amino acids and its derivatives in microorganisms and review the recent advances in bio-production of value-added chemicals using synthetic biology.
Applications of Synthetic Biology in the Research of Natural Product
WANG Li-ping, LUO Yun-zi
2017, 33(1): 35-47. doi:
10.13560/j.cnki.biotech.bull.1985.2017.01.004
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As an important source of clinical medicine and new drug candidate,natural product plays an essential role in therapeutic and pharmaceutical industry. However,traditional methods for natural product discovery constrain the development of new compounds at certain extent,and identifying and refactoring the biosynthetic pathways of natural products provided new thoughts for the discovery of new natural product. Currently,with the prosperous development of bioinformatics and synthetic biology technologies,a new era of natural product discovery and engineering can be foreseen. Here,we summarize recent advances on the strategies of genetic recombination and gene cluster regulation related to synthetic biology techniques,and discuss their applications and issues in studying natural products.
Optimization Strategies for Synthetic Biological Systems of Natural Products
KUANG Xue-jun, ZOU Li-qiu, SUN Chao, CHEN Shi-lin
2017, 33(1): 48-57. doi:
10.13560/j.cnki.biotech.bull.1985.2017.01.005
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Natural products have become an important source of new drugs. However,there are some deficiencies while producing complex natural products by classical organic synthesis,one measure to efficiently make up those deficiencies is to apply synthetic biology in the biosynthesis of natural product,i.e.,designing and reconstructing efficient biosynthetic pathways of target compounds,re-engineering it in host cells,and producing target compounds by fermentation. However,achieving the yield of target product to the industrial level through technology of synthetic biology is still very challenging though there have been advances in synthetic biology. Here,we review various pathway optimization strategies for the synthetic biological systems of natural products. Via optimization technologies of fine-tuning individual component,exogenous metabolic pathways,the chassis systems,and fermentation conditions,the synthetic biological system may be optimized,and the yield of target product may be maximized,thus providing continuous,stable and economical raw material supplies for the manufacture of complex natural products while source is rare,and promoting the research and development of new drugs similar to natural products
Application of Switch for Synthetic Biology in Metabolic Engineering
PANG Qing-xiao, LIANG Quan-feng, QI Qing-sheng
2017, 33(1): 58-64. doi:
10.13560/j.cnki.biotech.bull.1985.2017.01.006
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Translational switches for synthetic biology are mainly used to regulate gene expression in the field of metabolic engineering research. Traditional metabolic engineering regulates the expression of specific genes by using knockout and overexpression. However,the knockout of genes usually leads to the decrease in growth,therefore,we need to close and activate the expression of specific gene at proper time,and translational switche for synthetic biology is the key tool to solve such issue. At present,light-controlled switch,temperature-induced switch,toggle switch,and riboswitch are commonly used in metabolic engineering,of which the toggle switch and riboswitch present a great advantage in the dynamic regulation of gene expression. In this paper,several kinds of translational switches for synthetic biology and their applications in metabolic engineering are reviewed.
Research Progresses in the Synthetic Biology of Terpenoids
SUN Li-chao, LI Shu-ying, WANG Feng-zhong, XIN Feng-jiao
2017, 33(1): 64-75. doi:
10.13560/j.cnki.biotech.bull.1985.2017.01.007
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The terpenoids represent the largest class of natural products with biological activities of antitumor and anti-allergy,thus they have been widely applied in the area of food,cosmetics and medical health,presenting huge potential and broad market prospects. Recent years,researchers applied functional genomics and metabonomics approaches to deeply study the biosynthesis pathways of terpenoids,providing tons of data for their synthetic biology. The construction of engineered yeasts using synthetic biology enabled the efficient synthesis of multi-target terpenoids,and highly improved the overall production level. Thus,the synthetic biology approach is expected to be an efficient way of producing plant-derived terpenoids. First,we introduced the concept of synthetic biology,summarized the important functions and applications of plant-derived terpenoids,briefly reviewed the biosynthesis pathways,and concluded the alternative production ways. Then,we discussed the design strategies of synthetic biology for terpenoids thoroughly. Finally,we elaborated the advances on the biosynthetic biology of varied terpenes with common terpenes as the studied cases.
Advances on Applications of Synthetic Biology and Directed Evolution in Microbial Systems
GUO Yuan, ZHAO Zhong-lin
2017, 33(1): 76-82. doi:
10.13560/j.cnki.biotech.bull.1985.2017.01.008
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Nowadays,there are needs to engineer complex multi-genic traits in modern biotechnology applications;however,our capability to rationally engineer them is limited by the lack of knowledge on the genetic basis of complex phenotypes. Using synthetic biological measures,complex phenotypes can be engineered at the systems level,and via directed evolution strategies the whole biological system may be driven toward desired phenotypes without requiring the knowledge of the genetic basis of the targeted traits. The latest developments in the synthetic biology accelerate the directed evolution cycle,facilitating engineering of increasingly complex traits in biological systems. Herein,the recent advances on synthetic biology in directed evolution of complex traits in microbial systems are reviewed.
Ultra-high-throughput Screening System Based on Droplet Microfluidics and Its Applications in Synthetic Biology
MA Fu-qiang, YANG Guang-yu
2017, 33(1): 83-92. doi:
10.13560/j.cnki.biotech.bull.1985.2017.01.009
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The construction of a highly efficient synthetic biological system requires laborious screening work to optimize the performance of artificial system. Droplet microfluidic techniques miniaturize traditional screening systems,in a uniform pico-liter micro-reactors,the detection of enzymatic reactions and metabolites at single-cell level were measured and screened,thus it can screen genes/cells at a speed up to >108 events per day,which greatly improves the optimizing capability of catalytic parts and engineered strains. Herein,the technical backgrounds while applying droplet microfluidics in synthetic biology system are summarized,focusing on the fluorescence-coupling strategies in the detection of target enzyme reactions and metabolites. Moreover,recent progress on employing droplet microfluidics in the screening and engineering of catalytic parts and metabolic pathways is reviewed,the developing trends of this field are discussed.
The Development of Engineered Saccharomyces cerevisiae for Biomass Conversion
LIU He, ZHU Jia-qing, ZONG Qiu-jin, LI Bing-zhi, YUAN Ying-jin
2017, 33(1): 93-98. doi:
10.13560/j.cnki.biotech.bull.1985.2017.01.018
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With the increasing demand of oil,coal and other nonrenewable resources,it is urgent to develop the use of renewable and eco-friendly energy resources. The production of cellulosic ethanol from biomass has the advantage that the raw material is renewable and the cost is low. Saccharomyces cerevisiae has been most widely used for biomass conversion,with the characters of being easy for genetic engineering operation and high inhibitor tolerance. The two major challenges during the process of S. cerevisiae used for biomass conversion are the utilization of xylose and inhibitor tolerance. The major emphasis of this review will be the development of engineering Saccharomyces cerevisiae to improve inhibitor tolerance and speed up the transformation of xylose utilization.
Research Progress on the Synthesis of S-adenosyl-L-methionine in Microorganism
ZHAO Wei-jun, HUANG Lei, XU Zhi-nan
2017, 33(1): 99-105. doi:
10.13560/j.cnki.biotech.bull.1985.2017.01.010
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S-adenosyl-L-methionine(SAM)as an important metabolite exists in all cell types of living organisms,presents promising effects on liver diseases and depressive syndromes without any side effects,thus,it is greatly demanded in medical market. Recently,broad studies on biosynthesis of SAM have been conducted by scholars in the world. On the one hand,fermentation strategies based on traditionally producing SAM were further improved;on the other hand,different rational breeding programs were developed,such as fine tuning host’s metabolic network or screening strains by synthetic gene circuit. In this paper,progress in the research of S-adenosyl-L-methionine production is summarized,and the significances and issues while efficiently producing SAM by microorganism are discussed,aiming at providing the guides for improving SAM productivity at an industrial scale.
Research Progress for Genetic Modification of Butanol-producing Clostridia
ZHANG Chao, WANG Yi-qiang, WANG Qi-ye, HUANG Rui-chun, MI Xiao-qin
2017, 33(1): 106-113. doi:
10.13560/j.cnki.biotech.bull.1985.2017.01.011
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Butanol-producing Clostridia as butanol fermentation strains have been studied frequently in recent years. As a new renewable energy sources,butanol has obviously more advantages than ethanol. Therefore,it is of great significance to study gene modification of butanol-producing Clostridia. In this review,from three aspects of key genes,glycolytic pathway,and butanol tolerance,we introduced the latest research progress on gene modification of butanol-producing Clostridia. Meanwhile,we discussed the issues in the current research,and put forward suggestions on how to improve the butanol yield,aiming at providing some new ideas for the researchers.
Research Progresses on Riboswitches and Their Applications in Antimicrobials
SHENG Shu-yue, CHEN Yue, ZHANG Xing-mei, SHI Yu-sheng
2017, 33(1): 114-119. doi:
10.13560/j.cnki.biotech.bull.1985.2017.01.012
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Riboswitch is a class of RNA elements that regulate expressions of corresponding downstream genes and then control cells’ functions by binding with nucleic acids,amino acids,metal ions,derivatives of carbohydrates and coenzymes. Riboswitch has been a research focus in gene regulation in recent years and has been being used in mass screening bacterial strains,constructing new biosensors and as new target of antimicrobials. In this paper,recent research progresses on several important classes of riboswitches(such as purine riboswitch,lysine riboswitch,c-di-GMP riboswitch,glmS riboswitch,TPP riboswitch,FMN riboswitch,etc.)in targeting of antimicrobials are mainly reviewed.
Activity Comparison of the Artificial Hybrid Promoter with Its Native Promoter in Saccharomyces cerevisiae
TANG Rui-qi, XIONG Liang, BAI Feng-wu, ZHAO Xin-qing
2017, 33(1): 120-128. doi:
10.13560/j.cnki.biotech.bull.1985.2017.01.013
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The responses of promoter strengths of artificial TEF1 promoter(PaTEF1)and native promoter PTEF1 as well as PTDH3 were comprehensively compared. The strength of PaTEF1 was not always higher than PTEF1,but varied along with the genetic background of host,medium,and cell growth phase. Among the three investigated hosts,PaTEF1 showed the highest strength in BY4741,while the lowest in LX03. The strengths of PaTEF1 were 1.4-4.6 and 0.9-2.0 times of those of native promoters(PTEF1 and PTDH3)in YPD100 medium,respectively. The activities of PaTEF1 in YPE(5% and 7%)were 0.7-1.3 and 0.8-1.3 times of PTEF1 and PTDH3. The activity of PaTEF1 in YPE was 1.7-2.0 times of that in YPD100,whereas 2.7-7.1 and 1.3-3.4 times for PTEF1 and PTDH3,respectively. The activities of the promoters in YEP were higher than those in YPD100,however,the activity variation of artificial promoters was less than that of native promoter. In addition,the variation trends of promoter strengths from early-log to mid-log phase and from mid-log to stationary phase varied in different host strains.
Biosynthesis of Valerena-4,7(11)-diene in an Engineered Escherichia coli Strain
LIU Chang, YIN Hua, ZHUANG Yi-bin, LIU Tao
2017, 33(1): 129-134. doi:
10.13560/j.cnki.biotech.bull.1985.2017.01.014
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Valerena-4,7(11)-diene is a bicyclic sesquiterpene and shows particularly profound sedative,stress reducing and anxiolytic effects,and it could be used to treat attention deficit hyperactivity disorder(ADHD)and other mental illness. To achieve the de novo biosynthesis of valerena-4,7(11)-diene in Escherichia coli,eight genes of Saccharomyces cerevisiae involved in converting acetyl-CoA into farnesyl diphosphate(FPP)from mevalonate pathway and the valerena-4,7(11)-diene synthase(VoTPS)gene from Valeriana officinalis were co-overexpressed in BL21(DE3). Adopting the bipolar fermentation method of water phase and organic phase,the valerian-4,7(11)-diene was detected in organic phase by GC-MS,indicating that the synthesis of valerian-4,7(11)-diene was achieved initially in theE. coli. After primarily optimizing the IPTG concentration,the temperature of protein induction and choice of varied organic phases,the fermentation conditions were determined as follows:protein induced temperature was 20℃,the concentration of IPTG was 0.1 mmol/L,and organic phase was dodecane.
Biosynthesis of 3,4-dihydroxymandelic Acid in an Engineered Escherichia coli Strain
LI Xiao-lin, ZHOU Wei, ZHUANG Yi-bin, LU Fu-ping, YIN Hua
2017, 33(1): 135-140. doi:
10.13560/j.cnki.biotech.bull.1985.2017.01.015
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3,4-dihydroxymandelic acid,an important pharmaceutical and spices intermediate,possesses strong antioxidative and radical scavenging activities. To achieve the de novo biosynthesis of 3,4-dihydroxymandelic acid in Escherichia coli,the genes of hydroxymandelate synthase(HmaS)from Streptomyces coelicolor and 4-hydroxyphenylacetate 3-hydroxylase(HpaBC)from E. coli were cloned into pTrcHisB and overexpressed in an engineered E. coli strain MG1655/ΔA. After induction of IPTG and optimization of the inducing temperature,the yield of 3,4-dihydroxymandelic acid reached 240 mg/L in shake flask after 36 hours fermentation,laying a foundation for the large-scale fermentation production of 3,4-dihydroxymandelic acid .
Regulatory Effects of Gene sco1135 on the Sporulation and Secondary Metabolite Synthesis of Streptomyces coelicolor M145
MA Jun-xia, ZHANG Pei-pei, WANG Shi-li, CAO Guang-xiang,
2017, 33(1): 141-147. doi:
10.13560/j.cnki.biotech.bull.1985.2017.01.016
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This work aims to study the effects of deletion and mutation of gene sco1135 on the morphogenesis and secondary metabolism in Streptomyces coelicolor strain M145. Recombinant plasmid pSJ1135 was obtained by PCR-targeting. Gene deletion mutant of sco1135(△sco1135)was acquired by introducing the recombinant plasmid into the Streptomyces coelicolor M145 via conjugation and transformation,and the complemented strain △sco1135com was constructed using the vector pMS82. Using pMS82 as a control,the phenotypes of the parental wild strain(M145),mutant strain(△sco1135)and complemented strain(△sco1135com)were analyzed and observed with quantitative antibiotics. The results of phenotypic analysis and observation with quantitative antibiotics revealed that the sporulation in △sco1135 obviously delayed than that of the wild type M145 on YBP medium;while the actinorhodin(ACT)production in △sco1135 increased to 2-3 folds of that in M145. The transcriptions of some sporulation-related genes decreased by 50%-75% in △sco1135 at 48 h,while the transcriptional expressions of genes related to ACT in △sco1135 increased to 13-20 folds at 72 h,compared to the M145. Conclusively,sco1135 is involved in regulating the sporulation in M145 and also the production of secondary metabolite ACT.
Construction of 5-aminolevulinic Acid Synthesis Pathway and Optimization of Fermentation by Corynebacterium glutamicum
RAO De-ming, ZHANG Liang-cheng, CHEN Jiu-zhou, SUN De-hu, SUN Cun-min, ZHENG Xiao-mei, ZHENG Ping, DIAO Ai-po
2017, 33(1): 148-156. doi:
10.13560/j.cnki.biotech.bull.1985.2017.01.017
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5-aminolevulinic acid(ALA)is a natural non-protein amino acid and has been widely applied in agriculture and medicine. This study aimed to construct the systhesis pathway of C4 ALA in Corynebacterium glutamicum and optimize its fermentation process. First,efficient C4 ALA synthesis pathway was established in C. glutamicum by over-expressing the ALA synthase(ALAS)from Rhodopseudomonas palustris. Then,the ALA fermentation process with flask was optimized from four factors:fermentation medium,concentration of inducer,substrate concentration,and initial inoculum dosage. As results,the ALA yield of the strain 13032/pZWA1 over-expressing HemA was 1.41 g/L,up to 67.14 folds compared with the control strain. The optimal ALA fermentation condition was M9 media using yeast extract as nitrogen source with 5% inoculum size,0.1 mmol/L IPTG to induce the experession of HemA,and the concentration of glycine must be at 4 g/L. After optimization,ALA yield reached 3.28 g/L in a shaking flask and increased 132.62% than before. In conclusion,under the optimal fermentation condition,the ALA yield in a 5 L bioreactor fermentation was 10.08 g/L,which was the highest ALA yield by one-step fermentation of C. glutamicum reported so far.