Non-thermal plasma(NTP)is considered to be a promising technology for the removal of volatile organic compounds;however,its application is limited by low CO_(2) selectivity and undesirable by-products.To overcome thes...Non-thermal plasma(NTP)is considered to be a promising technology for the removal of volatile organic compounds;however,its application is limited by low CO_(2) selectivity and undesirable by-products.To overcome these issues,this paper discusses the degradation of chlorobenzene(CB)in systems of NTP coupled with catalysts,and the influence of catalyst locations in the NTP was investigated.In addition,the interaction between plasma and catalyst was also explored.The results indicated that the degradability of CB was remarkably improved through the combination of NTP with catalysts,and the formation of ozone was effectively inhibited.The degradation efficiency increased from 33.9%to 79.6%at 14 kV in the NTPcatalytic system,while the ozone concentration decreased from 437 to 237 mg m^(-3),and the degradation efficiency of in plasma catalysis(IPC)systems was superior to that of the post plasma catalysis system,while the inhibition ability of ozone exhibited an opposing trend.In the IPC system,the degradation efficiency was 87.7%at 14 k V,while the ozone concentration was151 mg m^(-3).Besides,the plasma did not destroy the pore structure and crystal structure of the catalyst,but affected the surface morphology and redox performance of the catalyst.Thus,NTP coupled catalytic system could improve the degradation performance of CB.Furthermore,the plasma discharge characteristics played a major role in the NTP synergistic catalytic degradation of CB.Finally,based on the experiment analysis results,the general reaction mechanism of CB degradation in an IPC reaction system was proposed.展开更多
As a fundamental tool in synthetic biology,promoters are pivotal in regulating gene expression,enabling precise genetic control and spurring innovation across diverse biotechnological applications.However,most advance...As a fundamental tool in synthetic biology,promoters are pivotal in regulating gene expression,enabling precise genetic control and spurring innovation across diverse biotechnological applications.However,most advances in engineered genetic systems rely on host-specific regulation of the genetic portion.With the burgeoning diversity of synthetic biology chassis cells,there emerges a pressing necessity to broaden the universal promoter toolkit spectrum,ensuring adaptability across various microbial chassis cells for enhanced applicability and custom-ization in the evolving landscape of synthetic biology.In this study,we analyzed and validated the primary structures of natural endogenous promoters from Escherichia coli,Bacillus subtilis,Corynebacterium glutamicum,Saccharomyces cerevisiae,and Pichia pastoris,and through strategic integration and rational modification of promoter motifs,we developed a series of cross-species promoters(P_(sh))with transcriptional activity in five strains(prokaryotic and eukaryotic).This series of cross species promoters can significantly expand the synthetic biology promoter toolkit while providing a foundation and inspiration for standardized development of universal components The combinatorial use of key elements from prokaryotic and eukaryotic promoters presented in this study represents a novel strategy that may offer new insights and methods for future advancements in promoter engineering.展开更多
Bacillus subtilis as the Gram-positive model bacterium has been widely used in synthetic biology and biotechnology while the regulatory RNA tools for B.subtilis are still not fully explored.Here,a bottom-up approach i...Bacillus subtilis as the Gram-positive model bacterium has been widely used in synthetic biology and biotechnology while the regulatory RNA tools for B.subtilis are still not fully explored.Here,a bottom-up approach is proposed for designing artificial trans-acting sRNAs.By engineering the intrinsic sRNA SR6,a minimized core scaffold structure consisting of an 8 bp stem,a 4 nt loop,and a 9 nt polyU tail was generated and proven to be sufficient for constructing sRNAs with strong repression activity(83%).Moreover,we demonstrate this artificial sRNA system functions well in an hfq-independent manner and also achieves strong repression efficiency in Escherichia coli(above 80%).A structure-based sRNA design principle was further developed for the automatic generation of custom sRNAs with this core scaffold but various sequences,which facilitates the manipulation and avoids structure disruption when fusing any base-pairing sequence.By applying these auto-designed sRNAs,we rapidly modified the cell morphology and biofilm formation,and regulated metabolic flux toward acetoin biosynthesis.This sRNA system with cross-species regulatory activities not only enriched the gene regulation toolkit in synthetic biology for B.subtilis and E.coli but also enhanced our understanding of trans-acting sRNAs.展开更多
Gene regulation by trans-acting small RNAs(sRNAs)has considerable advantages over other gene regulation strategies.However,synthetic sRNAs mainly take natural sRNAs(MicC or SgrS)as backbones and comprise three functio...Gene regulation by trans-acting small RNAs(sRNAs)has considerable advantages over other gene regulation strategies.However,synthetic sRNAs mainly take natural sRNAs(MicC or SgrS)as backbones and comprise three functional elements folding into two or more stem-loop structures:an mRNA base pairing region,an Hfq-binding structure,and a rho-independent terminator.Due to limited numbers of natural sRNAs and complicated backbone structures,synthetic sRNAs suffer from low activity programmability and poor structural modularity.Moreover,natural sRNA backbone sequences may increase the possibility of unwanted recombination.Here,we present a bottom-up approach for creating structure defined single-stem loop small non-coding RNAs(ssl-sRNAs),which contain a standardized scaffold of a 7 bp-stem-4 nt-loop-polyU-tail and a 24 nt basing pairing region covering the first eight codons.Particularly,ssl-sRNA requires no independent Hfq-binding structure,as the polyU tail fulfills the roles of binding Hfq.A thermodynamic-based scoring model and a web server sslRNAD(http://www.kangzlab.cn/)were developed for automated design of ssl-sRNAs with well-defined structures and programmable activities.ssl-sRNAs displayed weak polar effects when regulating polycistronic mRNAs.The ssl-sRNA designed by sslRNAD showed regulatory activities in both Escherichia coli and Bacillus subtilis.A streamlined workflow was developed for the construction of customized ssl-sRNA and ssl-sRNA libraries.As examples,the E.coli cell morphology was easily modified and new target genes of ergothioneine biosynthesis were quickly identified with ssl-sRNAs.ssl-sRNA and its designer sslRNAD enable researchers to rapidly design sRNAs for knocking down target genes.展开更多
Bacteria exhibit a rich repertoire of RNA molecules that intricately regulate gene expression at multiple hierarchical levels,including small RNAs(sRNAs),riboswitches,and antisense RNAs.Notably,the majority of these r...Bacteria exhibit a rich repertoire of RNA molecules that intricately regulate gene expression at multiple hierarchical levels,including small RNAs(sRNAs),riboswitches,and antisense RNAs.Notably,the majority of these regulatory RNAs lack or have limited protein-coding capacity but play pivotal roles in orchestrating gene expression by modulating transcription,post-transcription or translation processes.Leveraging and redesigning these regulatory RNA elements have emerged as pivotal strategies in the domains of metabolic engineering and synthetic biology.While previous investigations predominantly focused on delineating the roles of regulatory RNA in Gram-negative bacterial models such as Escherichia coli and Salmonella enterica,this review aims to summarize the mechanisms and functionalities of endogenous regulatory RNAs inherent to typical Gram-positive bacteria,notably Bacillus subtilis.Furthermore,we explore the engineering and practical applications of these regulatory RNA elements in the arena of synthetic biology,employing B.subtilis as a foundational chassis.展开更多
Escherichia coli is the most well-studied model prokaryote and has become an indispensable host for the biotech-nological production of proteins and biochemicals.In particular,the probiotic status of one E.coli strain...Escherichia coli is the most well-studied model prokaryote and has become an indispensable host for the biotech-nological production of proteins and biochemicals.In particular,the probiotic status of one E.coli strain,E.coli Nissle 1917(EcN)has helped it become a new favorite amongst synthetic biologists.To broaden its potential applications,here we assemble a comparative study on the genomes,transcriptomes,and metabolic properties of E.coli strains EcN,BL21(DE3),and MG1655.Comparative genomics data suggests that EcN possesses 1404 unique CDSs.In particular,EcN has additional iron transport systems which endow EcN with a higher tolerance to iron scarcity when compared to two other E.coli strains.EcN transcriptome data demonstrates that E.coli strains EcN,BL21(DE3),and MG1655 all have comparable activities of the central metabolic pathway,however only EcN inherits the arginine deiminase pathway.Additionally,we found that EcN displayed a lower expres-sion of ribosomal proteins compared to BL21(DE3)and MG1655.This comparative study on E.coli strains EcN,BL21(DE3),and MG1655 aims to provide a reference for further engineering EcN as a biotechnological tool.展开更多
基金supported by the National Key Research and Development Program of China(No.2018YFC1903100)Beijing Municipal Science and Technology Project Program(No.Z191100009119002)the State Environmental Protection Key Laboratory of Odor Pollution Control(No.20210504)。
文摘Non-thermal plasma(NTP)is considered to be a promising technology for the removal of volatile organic compounds;however,its application is limited by low CO_(2) selectivity and undesirable by-products.To overcome these issues,this paper discusses the degradation of chlorobenzene(CB)in systems of NTP coupled with catalysts,and the influence of catalyst locations in the NTP was investigated.In addition,the interaction between plasma and catalyst was also explored.The results indicated that the degradability of CB was remarkably improved through the combination of NTP with catalysts,and the formation of ozone was effectively inhibited.The degradation efficiency increased from 33.9%to 79.6%at 14 kV in the NTPcatalytic system,while the ozone concentration decreased from 437 to 237 mg m^(-3),and the degradation efficiency of in plasma catalysis(IPC)systems was superior to that of the post plasma catalysis system,while the inhibition ability of ozone exhibited an opposing trend.In the IPC system,the degradation efficiency was 87.7%at 14 k V,while the ozone concentration was151 mg m^(-3).Besides,the plasma did not destroy the pore structure and crystal structure of the catalyst,but affected the surface morphology and redox performance of the catalyst.Thus,NTP coupled catalytic system could improve the degradation performance of CB.Furthermore,the plasma discharge characteristics played a major role in the NTP synergistic catalytic degradation of CB.Finally,based on the experiment analysis results,the general reaction mechanism of CB degradation in an IPC reaction system was proposed.
基金supported by the National Key Research and Development Program of China(2021YFC2100800)the National Natural Science Foundation of China(32370066)+1 种基金the Fundamental Research Funds for the Central Universities(JUSRP622003)the National First-class Discipline Program of Light Industry Technology and Engineering(10152130122301801004).
文摘As a fundamental tool in synthetic biology,promoters are pivotal in regulating gene expression,enabling precise genetic control and spurring innovation across diverse biotechnological applications.However,most advances in engineered genetic systems rely on host-specific regulation of the genetic portion.With the burgeoning diversity of synthetic biology chassis cells,there emerges a pressing necessity to broaden the universal promoter toolkit spectrum,ensuring adaptability across various microbial chassis cells for enhanced applicability and custom-ization in the evolving landscape of synthetic biology.In this study,we analyzed and validated the primary structures of natural endogenous promoters from Escherichia coli,Bacillus subtilis,Corynebacterium glutamicum,Saccharomyces cerevisiae,and Pichia pastoris,and through strategic integration and rational modification of promoter motifs,we developed a series of cross-species promoters(P_(sh))with transcriptional activity in five strains(prokaryotic and eukaryotic).This series of cross species promoters can significantly expand the synthetic biology promoter toolkit while providing a foundation and inspiration for standardized development of universal components The combinatorial use of key elements from prokaryotic and eukaryotic promoters presented in this study represents a novel strategy that may offer new insights and methods for future advancements in promoter engineering.
基金supported by the National Natural Science Foundation of China (31970085)the National Key Research and Development Program of China (2021YFC2100800)the Jiangsu Province Natural Science Fund for Distinguished Young Scholars (BK20200025).
文摘Bacillus subtilis as the Gram-positive model bacterium has been widely used in synthetic biology and biotechnology while the regulatory RNA tools for B.subtilis are still not fully explored.Here,a bottom-up approach is proposed for designing artificial trans-acting sRNAs.By engineering the intrinsic sRNA SR6,a minimized core scaffold structure consisting of an 8 bp stem,a 4 nt loop,and a 9 nt polyU tail was generated and proven to be sufficient for constructing sRNAs with strong repression activity(83%).Moreover,we demonstrate this artificial sRNA system functions well in an hfq-independent manner and also achieves strong repression efficiency in Escherichia coli(above 80%).A structure-based sRNA design principle was further developed for the automatic generation of custom sRNAs with this core scaffold but various sequences,which facilitates the manipulation and avoids structure disruption when fusing any base-pairing sequence.By applying these auto-designed sRNAs,we rapidly modified the cell morphology and biofilm formation,and regulated metabolic flux toward acetoin biosynthesis.This sRNA system with cross-species regulatory activities not only enriched the gene regulation toolkit in synthetic biology for B.subtilis and E.coli but also enhanced our understanding of trans-acting sRNAs.
基金supported by the National Key Research and Development Program of China (2021YFC2100800)the National Natural Science Foundation of China (31970085)the Jiangsu Province Natural Science Fund for Distinguished Young Scholars (BK20200025).
文摘Gene regulation by trans-acting small RNAs(sRNAs)has considerable advantages over other gene regulation strategies.However,synthetic sRNAs mainly take natural sRNAs(MicC or SgrS)as backbones and comprise three functional elements folding into two or more stem-loop structures:an mRNA base pairing region,an Hfq-binding structure,and a rho-independent terminator.Due to limited numbers of natural sRNAs and complicated backbone structures,synthetic sRNAs suffer from low activity programmability and poor structural modularity.Moreover,natural sRNA backbone sequences may increase the possibility of unwanted recombination.Here,we present a bottom-up approach for creating structure defined single-stem loop small non-coding RNAs(ssl-sRNAs),which contain a standardized scaffold of a 7 bp-stem-4 nt-loop-polyU-tail and a 24 nt basing pairing region covering the first eight codons.Particularly,ssl-sRNA requires no independent Hfq-binding structure,as the polyU tail fulfills the roles of binding Hfq.A thermodynamic-based scoring model and a web server sslRNAD(http://www.kangzlab.cn/)were developed for automated design of ssl-sRNAs with well-defined structures and programmable activities.ssl-sRNAs displayed weak polar effects when regulating polycistronic mRNAs.The ssl-sRNA designed by sslRNAD showed regulatory activities in both Escherichia coli and Bacillus subtilis.A streamlined workflow was developed for the construction of customized ssl-sRNA and ssl-sRNA libraries.As examples,the E.coli cell morphology was easily modified and new target genes of ergothioneine biosynthesis were quickly identified with ssl-sRNAs.ssl-sRNA and its designer sslRNAD enable researchers to rapidly design sRNAs for knocking down target genes.
基金This work was supported by the National Natural Science Foundation of China(31970085 and 32000058)the National Key Research and Development Program of China(2021YFC2100800).
文摘Bacteria exhibit a rich repertoire of RNA molecules that intricately regulate gene expression at multiple hierarchical levels,including small RNAs(sRNAs),riboswitches,and antisense RNAs.Notably,the majority of these regulatory RNAs lack or have limited protein-coding capacity but play pivotal roles in orchestrating gene expression by modulating transcription,post-transcription or translation processes.Leveraging and redesigning these regulatory RNA elements have emerged as pivotal strategies in the domains of metabolic engineering and synthetic biology.While previous investigations predominantly focused on delineating the roles of regulatory RNA in Gram-negative bacterial models such as Escherichia coli and Salmonella enterica,this review aims to summarize the mechanisms and functionalities of endogenous regulatory RNAs inherent to typical Gram-positive bacteria,notably Bacillus subtilis.Furthermore,we explore the engineering and practical applications of these regulatory RNA elements in the arena of synthetic biology,employing B.subtilis as a foundational chassis.
基金supported by the National Key Re-search and Development Program of China(2021YFC2100800)the Jiangsu Province Natural Science Fund for Distinguished Young Schol-ars(BK20200025)+1 种基金a grant from the Key Technologies R&D Program of Jiangsu Province(BE2019630)the National First-class Discipline Program of Light Industry Technology and Engineering(LITE2018-16).
文摘Escherichia coli is the most well-studied model prokaryote and has become an indispensable host for the biotech-nological production of proteins and biochemicals.In particular,the probiotic status of one E.coli strain,E.coli Nissle 1917(EcN)has helped it become a new favorite amongst synthetic biologists.To broaden its potential applications,here we assemble a comparative study on the genomes,transcriptomes,and metabolic properties of E.coli strains EcN,BL21(DE3),and MG1655.Comparative genomics data suggests that EcN possesses 1404 unique CDSs.In particular,EcN has additional iron transport systems which endow EcN with a higher tolerance to iron scarcity when compared to two other E.coli strains.EcN transcriptome data demonstrates that E.coli strains EcN,BL21(DE3),and MG1655 all have comparable activities of the central metabolic pathway,however only EcN inherits the arginine deiminase pathway.Additionally,we found that EcN displayed a lower expres-sion of ribosomal proteins compared to BL21(DE3)and MG1655.This comparative study on E.coli strains EcN,BL21(DE3),and MG1655 aims to provide a reference for further engineering EcN as a biotechnological tool.
