“Living therapeutic carriers” present a promising avenue for cancer research, but it is still challenging to achieve uniform and durable distribution of payloads throughout the solid tumor owing to the tumor microenvironment heterogeneity. Herein, a living drug sprinkle biohybrid (YB1–HCNs) was constructed by hitching acid/enzyme-triggered detachable nanoparticles (HCNs) backpack on the surface of metabolic oligosaccharide-engineered oncolytic bacteria YB1. Along with the process of tumor penetration by bacterial hypoxia navigation, YB1–HCNs responsively and continuously release HCNs, achieving a uniform distribution of loaded agents throughout the tumor. Upon successive irradiation of laser and ultrasound (US), the HCNs can separately generate type II and type I ROS for superior sono–photodynamic therapy (SPDT), which enables HCNs to synergize with YB1 for a satisfactory therapeutic effect in both superficial normoxic and deep hypoxic regions of the tumor. After a single dose, this efficient combination realized 98.3% primary tumor inhibition rate and prolonged survival of mice for 90 days with no recurrence, further inducing a powerful immunological memory effect to completely suppress tumor rechallenge in cured mice. Such a bacterial hybridization vector enables optimization of the spatial distribution of YB1 and HCNs, providing an innovative strategy to maximize therapeutic outcomes and evoke durable antitumor immunity.

Bacterial immunotherapy holds promising cancer-fighting potential. However, unlocking its power requires a mechanistic understanding of how bacteria both evade antimicrobial immune defenses and stimulate antitumor immune responses within the tumor microenvironment (TME). Here, by harnessing an engineered Salmonella enterica strain with this dual proficiency, we unveiled a singular mechanism underlying. Specifically, the hysteretic nonlinearity of interleukin-10 receptor (IL-10R) expression drives tumor-infiltrated immune cells into a tumor-specific IL-10Rhi state. Bacteria leverage this to enhance tumor-associated macrophages producing IL-10, evade phagocytosis by tumor-associated neutrophils, and coincidently expand and stimulate the preexisting exhausted tumor-resident CD8+ T cells. This effective combination eliminated tumors, prevented recurrence, and inhibited metastasis across multiple tumor types. Analysis of human samples suggested that IL-10Rhi state might be a ubiquitous trait across human tumor types. Our study unveils the unsolved mechanism behind bacterial immunotherapy's dual challenge in solid tumors and provides a framework for intratumoral immunomodulation.

Broad discussion is needed to chart a path forward.

Synthetic biology faces major challenges in the rational design of complex living systems, necessitating a quantitative understanding of the principles that guide the emergence of functions from biological building blocks. Here, we propose quantitative synthetic biology as a new research paradigm, integrating quantitative biology, systems biology and synthetic biology.

The robust regulation of the cell cycle is critical for the survival and proliferation of bacteria. To gain a comprehensive understanding of the mechanisms regulating the bacterial cell cycle, it is essential to accurately quantify cell-cycle-related parameters and to uncover quantitative relationships. In this paper, we demonstrate that the quantification of cell size parameters using microscopic images can be influenced by software and by the parameter settings used. Remarkably, even if the consistent use of a particular software and specific parameter settings is maintained throughout a study, the type of software and the parameter settings can significantly impact the validation of quantitative relationships, such as the constant-initiation-mass hypothesis. Given these inherent characteristics of microscopic image-based quantification methods, it is recommended that conclusions be cross-validated using independent methods, especially when the conclusions are associated with cell size parameters that were obtained under different conditions. To this end, we presented a flexible workflow for simultaneously quantifying multiple bacterial cell-cycle-related parameters using microscope-independent methods.

Synthetic biology provides a new paradigm for life science research ("build to learn") and opens the future journey of biotechnology ("build to use"). Here, we discuss advances of various principles and technologies in the mainstream of the enabling technology of synthetic biology, including synthesis and assembly of a genome, DNA storage, gene editing, molecular evolution and de novo design of function proteins, cell and gene circuit engineering, cell-free synthetic biology, artificial intelligence (AI)-aided synthetic biology, as well as biofoundries. We also introduce the concept of quantitative synthetic biology, which is guiding synthetic biology towards increased accuracy and predictability or the real rational design. We conclude that synthetic biology will establish its disciplinary system with the iterative development of enabling technologies and the maturity of the core theory.

Alternative splicing (AS) plays significant roles in a multitude of fundamental biological activities. AS is prevalent in the testis, but the regulations of AS in spermatogenesis is only little explored. Here, we report that Serine/arginine-rich splicing factor 1 (SRSF1) plays critical roles in alternative splicing and male reproduction. Male germ cell-specific deletion of Srsf1 led to complete infertility by affecting spermatogenesis. Mechanistically, by combining RNA-seq data with LACE-seq data, we showed that SRSF1 affected the AS of Stra8 in a direct manner and Dazl, Dmc1, Mre11a, Syce2 and Rif1 in an indirect manner. Our findings demonstrate that SRSF1 has crucial functions in spermatogenesis and male fertility by regulating alternative splicing.

RNA splicing plays significant roles in fundamental biological activities. However, our knowledge about the roles of alternative splicing and underlying mechanisms during spermatogenesis is limited.

Current strategies to improve the throughput of continuous directed evolution technologies often involve complex mechanical fluid-controlling system or robotic platforms, which limits their popularization and application in general laboratories. Inspired by our previous study on bacterial range expansion, in this study, we report a system termed SPACE for rapid and extensively parallelizable evolution of biomolecules by introducing spatial dimensions into the landmark phage-assisted continuous evolution system. Specifically, M13 phages and chemotactic Escherichia coli cells were closely inoculated onto a semisolid agar. The phages came into contact with the expanding front of the bacterial range, and then comigrated with the bacteria. This system leverages competition over space, wherein evolutionary progress is closely associated with the production of spatial patterns, allowing the emergence of improved or new protein functions. In a prototypical problem, SPACE remarkably simplified the process and evolved the promoter recognition of T7 RNA polymerase (RNAP) to a library of 96 random sequences in parallel. These results establish SPACE as a simple, easy to implement, and massively parallelizable platform for continuous directed evolution in general laboratories.

Whole-genome recoding has been shown to enable nonstandard amino acids, biocontainment and viral resistance in bacteria. Here we take the first steps to extend this to human cells demonstrating exceptional base editing to convert TAG to TAA for 33 essential genes via a single transfection, and examine base-editing genome-wide (observing ~40 C-to-T off-target events in essential gene exons). We also introduce GRIT, a computational tool for recoding. This demonstrates the feasibility of recoding, and highly multiplex editing in mammalian cells.

Bacterial cancer therapy, which was first applied in the clinic in 1868, has regained attention owing to the recent progress made in synthetic biology. Considering their easily manipulated genomes, preferential accumulation in tumors, and penetration abilities, bacteria have shown great therapeutic potential in tumor treatment. During treatment, it has been found that bacteria in tumors lead to corresponding changes in the abundance as well as locations of a variety of cells and substances, especially immune cells, forming a unique distribution pattern. This has been suggested to contribute to the therapeutic effect of bacterial cancer therapy

Transcription termination is one of the least understood processes of gene expression. As the prototype model for transcription studies, the single-subunit T7 RNA polymerase (RNAP) is known to respond to two types of termination signals, but the mechanism underlying such termination, especially the specific elements of the polymerase involved, is still unclear, due to a lack of knowledge with respect to the structure of the termination complex. Here we applied phage-assisted continuous evolution to obtain variants of T7 RNAP that can bypass the typical class I T7 terminator with stem-loop structure. Through in vivo selection and in vitro characterization, we discovered a single mutation (S43Y) that significantly decreased the termination efficiency of T7 RNAP at all transcription terminators tested. Coincidently, the S43Y mutation almost eliminates the RNA-dependent RNAP (RdRp) activity of T7 RNAP without impeding the major DNA-dependent RNAP (DdRp) activity of the enzyme. S43 is located in a hinge region and regulates the transformation between transcription initiation and elongation of T7 RNAP. Steady-state kinetics analysis and an RNA binding assay indicate that the S43Y mutation increases the transcription efficiency while weakening RNA binding of the enzyme. As an enzymatic reagent for in vitro transcription, the T7 RNAP S43Y mutant reduces the undesired termination in run-off RNA synthesis and produces RNA with higher terminal homogeneity.

Microbes in nature often live in dense and diverse communities exhibiting a variety of spatial structures. Microbial range expansion is a universal ecological process that enables populations to form spatial patterns. It can be driven by both passive and active processes, for example, mechanical forces from cell growth and bacterial motility. In this review, we provide a taste of recent creative and sophisticated efforts being made to address basic questions in spatial ecology and pattern formation during range expansion. We especially highlight the role of motility to shape community structures, and discuss the research challenges and future directions.

The engineered "obligate" anaerobic Salmonella typhimurium strain YB1 shows a prominent ability to repress tumor growth and metastasis, which has great potential as a novel cancer immunotherapy. However, the antitumor mechanism of YB1 remains unelucidated. To resolve the proteome dynamics induced by the engineered bacteria, we applied tumor temporal proteome profiling on murine bladder tumors after intravenous injection of either YB1 or PBS as a negative control. Our data suggests that during the two weeks treatment of YB1 injections, the cured tumors experienced three distinct phases of the immune response. Two days after injection, the innate immune response was activated, particularly the complement and blood coagulation pathways. In the meantime, the phagocytosis was initiated. The professional phagocytes such as macrophages and neutrophils were recruited, especially the infiltration of iNOS+ and CD68+ cells was enhanced. Seven days after injection, substantial amount of T cells was observed at the invasion margin of the tumor. As a result, the tumor shrunk significantly. Overall, the temporal proteome profiling can systematically reveal the YB1 induced immune responses in tumor, showing great promise for elucidating the mechanism of bacteria-mediated cancer immunotherapy.

Synthetic biology research and technology translation has garnered increasing interest from the governments and private investors in Asia, where the technology has great potential in driving a sustainable bio-based economy. This Perspective reviews the latest developments in the key enabling technologies of synthetic biology and its application in bio-manufacturing, medicine, food and agriculture in Asia. Asia-centric strengths in synthetic biology to grow the bio-based economy, such as advances in genome editing and the presence of biofoundries combined with the availability of natural resources and vast markets, are also highlighted. The potential barriers to the sustainable development of the field, including inadequate infrastructure and policies, with suggestions to overcome these by building public-private partnerships, more effective multi-lateral collaborations and well-developed governance framework, are presented. Finally, the roles of technology, education and regulation in mitigating potential biosecurity risks are examined. Through these discussions, stakeholders from different groups, including academia, industry and government, are expectantly better positioned to contribute towards the establishment of innovation and bio-economy hubs in Asia.

Bacterial cancer therapy, which was first applied in the clinic in 1868, has regained attention owing to the recent progress made in synthetic biology. Considering their easily manipulated genomes, preferential accumulation in tumors, and penetration abilities, bacteria have shown great therapeutic potential in tumor treatment. During treatment, it has been found that bacteria in tumors lead to corresponding changes in the abundance as well as locations of a variety of cells and substances, especially immune cells, forming a unique distribution pattern. This has been suggested to contribute to the therapeutic effect of bacterial cancer therapy

Self-organization is a prerequisite of biological complexity. At the population level, it amounts to spontaneously sorting differ ent individuals through space and time. Here, we reveal a simple mechanism by which different populations of motile cells can self-organize through a reciprocal control of their motilities. We first show how the reciprocal activation of motility between two populations of engineered Escherichia coli makes an initially mixed population of cells segregate, leading to out-of-phase population oscillations without the need of any preexisting positional or orientational cues. By redesigning the interaction, the original segregation between the two populations can be turned into co-localization. We account for this self-organization using a theoretical model that shows the reciprocal control of motility to be a robust and versatile self-organization pathway in multi-component systems. We finally show how our theoretical and experimental results can be generalized to three interacting bacterial populations.

Growth laws emerging from studies of cell populations provide essential constraints on the global mechanisms that coordinate cell growth1-3. The foundation of bacterial cell cycle studies relies on two interconnected dogmas that were proposed more than 50 years ago-the Schaechter-Maaloe-Kjeldgaard growth law that relates cell mass to growth rate1 and Donachie's hypothesis of a growth-rate-independent initiation mass4. These dogmas spurred many efforts to understand their molecular bases and physiological consequences5-14. Although they are generally accepted in the fast-growth regime, that is, for doubling times below 1 h, extension of these dogmas to the slow-growth regime has not been consistently achieved. Here, through a quantitative physiological study of Escherichia coli cell cycles over an extensive range of growth rates, we report that neither dogma holds in either the slow- or fast-growth regime. In their stead, linear relations between the cell mass and the rate of chromosome replication-segregation were found across the range of growth rates. These relations led us to propose an integral-threshold model in which the cell cycle is controlled by a licensing process, the rate of which is related in a simple way to chromosomal dynamics. These results provide a quantitative basis for predictive understanding of cell growth-cell cycle relationships.

The ability of a species to colonize newly available habitats is crucial to its overall fitness1-3. In general, motility and fast expansion are expected to be beneficial for colonization and hence for the fitness of an organism4-7. Here we apply an evolution protocol to investigate phenotypical requirements for colonizing habitats of different sizes during range expansion by chemotaxing bacteria8. Contrary to the intuitive expectation that faster is better, we show that there is an optimal expansion speed for a given habitat size. Our analysis showed that this effect arises from interactions among pioneering cells at the front of the expanding population, and revealed a simple, evolutionarily stable strategy for colonizing a habitat of a specific size: to expand at a speed given by the product of the growth rate and the habitat size. These results illustrate stability-to-invasion as a powerful principle for the selection of phenotypes in complex ecological processes.

Biofoundries provide an integrated infrastructure to enable the rapid design, construction, and testing of genetically reprogrammed organisms for biotechnology applications and research. Many biofoundries are being built and a Global Biofoundry Alliance has recently been established to coordinate activities worldwide.

Bacillary, Gram-negative bacteria grow by elongation with no discernible change in width, but during faster growth in richer media the cells are also wider. The mechanism regulating the change in cell width W during transitions from slow to fast growth is a fundamental, unanswered question in molecular biology. The value of W that changes in the divisome and during the division process only, is related to the nucleoid complexity, determined by the rates of growth and of chromosome replication; the former is manipulated by nutritional conditions and the latter-by thymine limitation of thyA mutants. Such spatio-temporal regulation is supported by existence of a minimal possible distance between successive replisomes, so-called eclipse that limits the number of replisomes to a maximum. Breaching this limit by slowing replication in fast growing cells results in maximal nucleoid complexity that is associated with maximum cell width, supporting the notion of Nucleoid-to-Divisome signal transmission. Physical signal(s) may be delivered from the nucleoid to assemble the divisome and to fix the value of W in the nascent cell pole.

Besides genome editing, CRISPR-Cas12a has recently been used for DNA detection applications with attomolar sensitivity but, to our knowledge, it has not been used for the detection of small molecules. Bacterial allosteric transcription factors (aTFs) have evolved to sense and respond sensitively to a variety of small molecules to benefit bacterial survival. By combining the single-stranded DNA cleavage ability of CRISPR-Cas12a and the competitive binding activities of aTFs for small molecules and double-stranded DNA, here we develop a simple, supersensitive, fast and high-throughput platform for the detection of small molecules, designated CaT-SMelor (CRISPR-Cas12a- and aTF-mediated small molecule detector). CaT-SMelor is successfully evaluated by detecting nanomolar levels of various small molecules, including uric acid and p-hydroxybenzoic acid among their structurally similar analogues. We also demonstrate that our CaT-SMelor directly measured the uric acid concentration in clinical human blood samples, indicating a great potential of CaT-SMelor in the detection of small molecules.

Bacteria-driven drug-delivery systems have attracted great attention for their enhanced therapeutic specificity and efficacy in cancer treatment. YB1, a particularly attractive genetically modified safe Salmonella Typhimurium strain, is known to penetrate hypoxic tumor cores with its self-driven properties while remarkably avoiding damage to normal tissues. Herein, nanophotosensitizers (indocyanine green (ICG)-loaded nanoparticles, INPs) were covalently attached to the surface of YB1 with amide bonds to develop a biotic/abiotic cross-linked system (YB1-INPs) for tumor precision therapy. YB1 microswimmer retained its viability after efficiently linking with INPs. This YB1-INPs treatment strategy demonstrated specific hypoxia targeting to solid tumors, perfect photothermal conversion, and efficient fluorescence (FL) imaging properties. Benefited from the combined contribution of tumor tissue destruction and the bacteria-attracting nutrients generation after photothermal treatment, the bioaccumulation of YB1-INPs was significantly improved 14-fold compared to no photothermal intervention. Furthermore, YB1-INPs pervaded throughout the large solid tumor (≥500 mm3). Under near-infrared (NIR) laser irradiation, YB1-INPs exhibited a dependable and highly efficient photothermal killing ability for eradicating the large solid tumor without relapse. This strategy of bacteria-driven hypoxia-targeting delivery has a great value for large solid tumors therapy with low toxicity and high efficiency.

Conventional techniques to synchronize bacterial cells often require manual manipulations and lengthy incubation lacking precise temporal control. An automated microfluidic device was recently developed to overcome these limitations. However, it exploits the stalk property of Caulobacter crescentus that undergoes asymmetric stalked and swarmer cell cycle stages and is therefore restricted to this species. To address this shortcoming, we have engineered Escherichia coli cells to adhere to microchannel walls via a synthetic and inducible "stalk". The pole of E. coli is capped by magnetic fluorescent nanoparticles via a polar-localized outer membrane protein. A mass of cells is immobilized in a microfluidic chamber by an externally applied magnetic field. Daughter cells are formed without the induced stalk and hence are flushed out, yielding a synchronous population of "baby" cells. The stalks can be tracked by GFP and nanoparticle fluorescence; no fluorescence signal is detected in the eluted cell population, indicating that it consists solely of daughters. The collected daughter cells display superb synchrony. The results demonstrate a new on-chip method to synchronize the model bacterium E. coli and likely other bacterial species, and also foster the application of synthetic biology to the study of the bacterial cell cycle.

Cellulose is the most abundant organic polymer mainly produced by plants in nature. It is insoluble and highly resistant to enzymatic hydrolysis. Cellulolytic microorganisms that are capable of producing a battery of related enzymes play an important role in recycling cellulose-rich plant biomass. Effective cellulose degradation by multiple synergic microorganisms has been observed within a defined microbial consortium in the lab culture. Metagenomic analysis may enable us to understand how microbes cooperate in cellulose degradation in a more complex microbial free-living ecosystem in nature.

Active matter comprises individual units that convert energy into mechanical motion. In many examples, such as bacterial systems and biofilament assays, constituent units are elongated and can give rise to local nematic orientational order. Such "active nematics" systems have attracted much attention from both theorists and experimentalists. However, despite intense research efforts, data-driven quantitative modeling has not been achieved, a situation mainly due to the lack of systematic experimental data and to the large number of parameters of current models. Here, we introduce an active nematics system made of swarming filamentous bacteria. We simultaneously measure orientation and velocity fields and show that the complex spatiotemporal dynamics of our system can be quantitatively reproduced by a type of microscopic model for active suspensions whose important parameters are all estimated from comprehensive experimental data. This provides unprecedented access to key effective parameters and mechanisms governing active nematics. Our approach is applicable to different types of dense suspensions and shows a path toward more quantitative active matter research.

As an alternative approach against multidrug-resistant bacterial infections, phages are now being increasingly investigated as effective therapeutic agents. Here, aiming to design an efficient phage cocktail against Aeromonas salmonicida infections, we isolated and characterized five lytic A. salmonicida phages, AS-szw, AS-yj, AS-zj, AS-sw, and AS-gz. The results of morphological and genomic analysis suggested that all these phages are affiliated to the T4virus genus of the Caudovirales order. Their heterogeneous lytic capacities against A. salmonicida strains were demonstrated by experiments. A series of phage cocktails were prepared and investigated in vitro. We observed that the cocktail combining AS-gz and AS-yj showed significantly higher antimicrobial activity than other cocktails and individual phages. Given the divergent genomes between the phages AS-yj and AS-gz, our results highlight that the heterogeneous mechanisms that phages use to infect their hosts likely lead to phage synergy in killing the host. Conclusively, our study described a strategy to develop an effective and promising phage cocktail as a therapeutic agent to combat A. salmonicida infections, and thereby to control the outbreak of relevant fish diseases. Our study suggests that in vitro investigations into phages are prerequisite to obtain satisfying phage cocktails prior to application in practice.

Substantial efforts have been made to link the gut bacterial community to many complex human diseases. Nevertheless, the gut phages are often neglected.

Cell size of bacteria M is related to 3 temporal parameters: chromosome replication time C,periodfrom replication-termination to subsequent division D, and doubling time t. Steady-state, bacillary cells grow exponentially by extending length L only, but their constant width W is larger at shorter t‘sorlongerC's, in proportion to the number of chromosome replication positions n (¼ C/t), at least in Escherichia coli and Salmonella typhimurium. Extending C by thymine limitation of fast-growing thyA mutants result in continuous increase of M,associatedwithrisingW, up to a limit before branching. A set of such puzzling observations is qualitatively consistent with the view that the actual cell mass (or volume) at the time of replication-initiation Mi (or Vi), usually relatively constant in growth at varying t0s, rises with time under thymine limitation of fast-growing, thymine-requiring E. coli strains. The hypothesis will be tested that presumes existence of a minimal distance lmin between successive moving replisomes, translated into the timeneededforareplisometoreachlminbeforeanewreplication-initiationatoriCisallowed,termedEclipse E. Preliminaryanalysisofcurrentlyavailabledataisinconsistent withaconstantEunderallconditions,hence otherexplanationsandwaystotestthemareproposedinanattempttoelucidatetheseandotherresults.The complex hypothesis takes into account much of what is currently known about Bacterial Physiology: the relationships between cell dimensions, growth and cycle parameters, particularly nucleoid structure, repli cationandposition,andthemodeofpeptidoglycanbiosynthesis.Furtherexperimentsarementionedthatare necessary to test the discussed ideas and hypotheses.

Quantitative biology is dedicated to taking advantage of quantitative reasoning and advanced engineering technologies to make biology more predictable. Microfluidics, as an emerging technique, provides new approaches to precisely control fluidic conditions on small scales and collect data in high-throughput and quantitative manners. In this review, the authors present the relevant applications of microfluidics to quantitative biology based on two major categories (channel-based microfluidics and droplet-based microfluidics), and their typical features. We also envision some other microfluidic techniques that may not be employed in quantitative biology right now, but have great potential in the near future.

Aims To demonstrate a nonempirical workflow to select host‐specific probiotics for aquaculture industry. Methods and Results Using both culture‐dependent and culture‐independent methods, we have systematically investigated, for the first time, the gut microbiota of twelve subtropical aquatic animal species. We found that the diversity, abundance and distribution of gut micro‐organisms of these animals were host‐specific and that lactic acid bacteria (LAB) were predominant among the indigenous probiotic microbes. Using culturing method, we isolated and characterized ninety‐eight LAB strains; however, only a few strains was representative of the dominant LAB OTUs recovered by culture‐independent analysis. Conclusions Two cultured LAB strains, Enterococcus faecalis LS1‐2 and Enterococcus faecium Z1‐2, capturing the major LAB OTUs in the sequencing data set of the most animal samples and showing significant antimicrobial activities against shrimp pathogens, were suggested to be the candidates of shrimp probiotics. Significance and Impact of Study Disease outbreak and the consequential abuse of antibiotics have been the constraints to the aquaculture industry. However, the selection of probiotic bacteria is currently still an empirical process due to our limited knowledge on the gastrointestinal microbiota of aquatic organisms. Our study points to a nonempirical selection process by which host‐specific probiotics can be developed.

Background The prediction of the prokaryotic promoter strength based on its sequence is of great importance not only in the fundamental research of life sciences but also in the applied aspect of synthetic biology. Much advance has been made to build quantitative models for strength prediction, especially the introduction of machine learning methods such as artificial neural network (ANN) has significantly improve the prediction accuracy. As one of the most important machine learning methods, support vector machine (SVM) is more powerful to learn knowledge from small sample dataset and thus supposed to work in this problem. Methods To confirm this, we constructed SVM based models to quantitatively predict the promoter strength. A library of 100 promoter sequences and strength values was randomly divided into two datasets, including a training set (⩾10 sequences) for model training and a test set (⩾10 sequences) for model test. Results The results indicate that the prediction performance increases with an increase of the size of training set, and the best performance was achieved at the size of 90 sequences. After optimization of the model parameters, a high-performance model was finally trained, with a high squared correlation coefficient for fitting the training set (R2 > 0.99) and the test set (R2 > 0.98), both of which are better than that of ANN obtained by our previous work. Conclusions Our results demonstrate the SVM-based models can be employed for the quantitative prediction of promoter strength.

Synthetic biology is an emerging interdisciplinary research field. By designing and constructing new or re-designing the existing natural systems, it confers them novel functions, which do not exist in nature. Owing to the predictability and controllability, synthetic biology attracts more and more interest from biologists, physicists, and engineers. Synthetic biology approaches not only can be widely used for biotechnological applications but also can be used to study complex biological systems to address fundamental questions. Here, we reviewed the recent studies following the concept of "build-to-understand", particularly, the studies to understand intracellular network structure, cell physiology, the behavior of multicellular populations, and ecosystems.

Long non-coding RNAs (lncRNAs) are prominently associated with chromosomes in an ever-increasing diversity of roles. To provide further insight into the potential nature of these associations, we have explored, for the first time, the interaction of long single-stranded (ss) RNAs with cognate homologous double-stranded (ds) DNA in vitro Using magnetic tweezers, we measured the effects of ssRNA on force extension curves for dsDNA. We observe that the presence of ssRNA impedes the extension of dsDNA, specifically at low forces, dependent on homology between the RNA and DNA species, and dependent on ssRNA lengths (≥1 kb). The observed effect also depends on the concentration of ssRNA and is abolished by overstretching of the dsDNA. These findings show that significant homologous contacts can occur between long ssRNA and dsDNA in the absence of protein and that these contacts alter the mechanical properties of the dsDNA. We propose that long ssRNA interacts paranemically with long dsDNA via periodic short homologous interactions, e.g. mediated by RNA/DNA triplex-formation, and that dsDNA extension is impeded by formation of RNA secondary structure in the intervening unbound regions. Analogous interactions in vivo would permit lncRNAs to mediate the juxtaposition of two or more DNA regions on the same or different chromosomes.

Bacteria tightly regulate and coordinate the various events in their cell cycles to duplicate themselves accurately and to control their cell sizes. Growth of Escherichia coli, in particular, follows a relation known as Schaechter's growth law. This law says that the average cell volume scales exponentially with growth rate, with a scaling exponent equal to the time from initiation of a round of DNA replication to the cell division at which the corresponding sister chromosomes segregate. Here, we sought to test the robustness of the growth law to systematic perturbations in cell dimensions achieved by varying the expression levels of mreB and ftsZ We found that decreasing the mreB level resulted in increased cell width, with little change in cell length, whereas decreasing the ftsZ level resulted in increased cell length. Furthermore, the time from replication termination to cell division increased with the perturbed dimension in both cases. Moreover, the growth law remained valid over a range of growth conditions and dimension perturbations. The growth law can be quantitatively interpreted as a consequence of a tight coupling of cell division to replication initiation. Thus, its robustness to perturbations in cell dimensions strongly supports models in which the timing of replication initiation governs that of cell division, and cell volume is the key phenomenological variable governing the timing of replication initiation. These conclusions are discussed in the context of our recently proposed "adder-per-origin" model, in which cells add a constant volume per origin between initiations and divide a constant time after initiation.

Synthetic biology is an interdisciplinary research field, combining disciplines such as life sciences, engineering, physics, and chemistry. Through designing and constructing new biological elements, functions, and systems, it aims to build controllable circuits, biological logics, and production systems, which do not exist in nature. Its potential application areas include medicine, environmental remediation, cancer therapy, and development of new pesticides, biofuels, and new materials. Synthetic biology has garnered the interest of not only scientists and engineers but also sociologists, economists, and philosophers. As currently used for this discipline, “Synthetic biology” is a broad concept, which indicates its multiple goals. What is synthetic biology? What is the scientific methodology and the philosophical implication? These issues will be discussed in this review.

生命从何而来? 生物进化的原理和分子机制是什么? 生物如何组装具有特定结构的分子和细胞, 又如何从一个细胞生长发育为一个有规则结构的生物体? 这些古老的生物学基本问题至今仍然蒙着神秘的面纱. 在过去几十年中, 合成生物学这门新兴交叉学科融合了生命科学、工程学、物理学与化学等诸多学科中的内容, 旨在通过设计和建造新的生物元件、功能和系统, 以创建在自然界中并不存在的可控方式、生物逻辑和生命系统. 合成生物学的出现或许能够克服此前的技术障碍, 在回答生命起源、进化、结构与功能等问题上提供新的有趣的观点; 此外, 它也可能改变对生命已被广泛接受的定义, 从而挑战认知生命的方式.

Acinetobacter sp. strain YZS-X1-1 was isolated from freshwater pond sludge in China. Here, we present the draft genome of strain YZS-X1-1, which consists of 3,278,660 bases, with a G+C content of 42.1%.

Engineered bacteria in which motility is reduced by local cell density generate periodic stripes of high and low density when spotted on agar plates. We study theoretically the origin and mechanism of this process in a kinetic model that includes growth and density-suppressed motility of the cells. The spreading of a region of immotile cells into an initially cell-free region is analyzed. From the calculated front profile we provide an analytic ansatz to determine the phase boundary between the stripe and the no-stripe phases. The influence of various parameters on the phase boundary is discussed.

The principles and molecular mechanisms underlying biological pattern formation are difficult to elucidate in most cases due to the overwhelming physiologic complexity associated with the natural context. The understanding of a particular mechanism, not to speak of underlying universal principles, is difficult due to the diversity and uncertainty of the biological systems. Although current genetic and biochemical approaches have greatly advanced our understanding of pattern formation, the progress mainly relies on experimental phenotypes obtained from time-consuming studies of gain or loss of function mutants. It is prevailingly considered that synthetic biology will come to the application age, but more importantly synthetic biology can be used to understand the life. Using periodic stripe pattern formation as a paradigm, we discuss how to apply synthetic biology in understanding biological pattern formation and hereafter foster the applications like tissue engineering.

Periodic stripe patterns are ubiquitous in living organisms, yet the underlying developmental processes are complex and difficult to disentangle. We describe a synthetic genetic circuit that couples cell density and motility. This system enabled programmed Escherichia coli cells to form periodic stripes of high and low cell densities sequentially and autonomously. Theoretical and experimental analyses reveal that the spatial structure arises from a recurrent aggregation process at the front of the continuously expanding cell population. The number of stripes formed could be tuned by modulating the basal expression of a single gene. The results establish motility control as a simple route to establishing recurrent structures without requiring an extrinsic pacemaker.

Alcanivorax dieselolei strain B-5 is a marine bacterium that can utilize a broad range of n-alkanes (C(5) -C(36) ) as sole carbon source. However, the mechanisms responsible for this trait remain to be established. Here we report on the characterization of four alkane hydroxylases from A. dieselolei, including two homologues of AlkB (AlkB1 and AlkB2), a CYP153 homologue (P450), as well as an AlmA-like (AlmA) alkane hydroxylase. Heterologous expression of alkB1, alkB2, p450 and almA in Pseudomonas putida GPo12 (pGEc47ΔB) or P. fluorescens KOB2Δ1 verified their functions in alkane oxidation. Quantitative real-time RT-PCR analysis showed that these genes could be induced by alkanes ranging from C(8) to C(36) . Notably, the expression of the p450 and almA genes was only upregulated in the presence of medium-chain (C(8) -C(16) ) or long-chain (C(22) -C(36) ) n-alkanes, respectively; while alkB1 and alkB2 responded to both medium- and long-chain n-alkanes (C(12) -C(26) ). Moreover, branched alkanes (pristane and phytane) significantly elevated alkB1 and almA expression levels. Our findings demonstrate that the multiple alkane hydroxylase systems ensure the utilization of substrates of a broad chain length range.

Targeted gene repair directed by single-stranded oligodeoxynucleotides (ssODNs) offers a promising tool for biotechnology and gene therapy. However, the methodology is currently limited by its low frequency of repair events, variability, and low viability of "corrected" cells. In this study, we showed that during ssODN-mediated gene repair reaction, a significant population of corrected cells failed to divide, and were much more prone to undergo apoptosis, as marked by processing of caspases and PARP-1. In addition, we found that apoptotic cell death triggered by ssODN-mediated gene repair was largely independent of the ATM/ATR kinase. Furthermore, we examined the potential involvement of the mismatch repair (MMR) proteins in this "correction reaction-induced" cell death. Result showed that while defective MMR greatly enhanced the efficiency of gene correction, compromising the MMR system did not yield any viable corrected clone, indicating that the MMR machinery, although plays a critical role in determining ssODN-directed repair, was not involved in the observed cellular genotoxic responses.

Two bacterial strains, M-5T and WP0211T, were isolated from the surface water of a waste-oil pool in a coastal dock and from a deep-sea sediment sample from the West Pacific Ocean, respectively. Analysis of 16S rRNA gene sequences indicated that both strains belonged to the class Alphaproteobacteria and were closely related to Thalassospira lucentensis (96.1 and 96.2 %, gene sequence similarity, respectively). Based on the results of physiological and biochemical tests, as well as DNA-DNA hybridization experiments, it is suggested that these isolates represent two novel species of the genus Thalassospira. Various traits allow both novel strains to be differentiated from Thalassospira lucentensis, including oxygen requirement, nitrate reduction and denitrification abilities and major fatty acid profiles, as well as their ability to utilize six different carbon sources. Furthermore, the novel strains may be readily distinguished from each other by differences in their motility, flagellation, growth at 4 degrees C and 40 degrees C, their ability to hydrolyse Tween 40 and Tween 80, their utilization of 19 different carbon sources and by quantitative differences in their fatty acid contents. It is proposed that the isolates represent two novel species for which the names Thalassospira xiamenensis sp. nov. (type strain, M-5T=DSM 17429T=CGMCC 1.3998T) and Thalassospira profundimaris sp. nov. (type strain, WP0211T=DSM 17430T=CGMCC 1.3997T) are proposed.

Salvianolic acid B (Sal B) is one of the most bioactive components of Salvia miltiorrhiza, a traditional Chinese herbal medicine that has been commonly used for prevention and treatment of cerebrovascular disorders. However, the mechanism responsible for such protective effects remains largely unknown. It has been considered that cerebral endothelium apoptosis caused by reactive oxygen species including hydrogen peroxide (H(2)O(2)) is implicated in the pathogenesis of cerebrovascular disorders.

Two bacterial strains, B-5(T) and NO1A, were isolated from the surface water of the Bohai Sea and deep-sea sediment of the east Pacific Ocean, respectively. Both strains were halophilic, aerobic, Gram-negative, non-spore-forming, catalase- and oxidase-positive motile rods. They grew on a restricted spectrum of organic compounds, including some organic acids and alkanes. On the basis of 16S rRNA gene sequence similarity, strains B-5(T) and NO1A were shown to belong to the gamma-Proteobacteria. Highest similarity values were found with Alcanivorax venustensis (95.2 %), Alcanivorax jadensis (94.6 %) and Alcanivorax borkumensis (94.1 %). Principal fatty acids of both strains were C(16 : 0), C(16 : 1)omega7c and C(18 : 1)omega7c. The chemotaxonomically characteristic fatty acid C(19 : 0) cyclo omega8c was also detected. On the basis of the above, together with results of physiological and biochemical tests, DNA-DNA hybridization, comparisons of 16S-23S internal transcribed spacer sequences and comparisons of the partial deduced amino acid sequence of alkane hydroxylase, both strains were affiliated to the genus Alcanivorax but were differentiated from recognized Alcanivorax species. Therefore, a novel species, Alcanivorax dieselolei sp. nov., represented by strains B-5(T) and NO1A is proposed, with the type strain B-5(T) (=DSM 16502(T)=CGMCC 1.3690(T)).