Chromosome recombination and modification by LoxP-mediated evolution in Vibrio natriegens using CRISPR-associated transposases.

Chromosome rearrangement by LoxP-mediated evolution has emerged as a powerful approach to studying how chromosome architecture impacts phenotypes. However, it relies on the in vitro synthesis of artificial chromosomes. The recently reported CRISPR-associated transposases (CASTs) held great promise for the efficient insertion of abundant LoxP sites directly onto the genome of wild-type strains. In this study, with the fastest-growing bacterium Vibrio natrigens (V. natriegens) as an object, a multiplex genome integration tool derived from CASTs was employed to achieve the insertion of cargo genes at eight specific genomic loci within 2 days. Next, we introduced 30 LoxP sites onto chromosome 2 (Chr2) of V. natriegens. Rigorously induced Cre recombinase was used to demonstrate Chromosome Rearrangement and Modification by LoxP-mediated Evolution (CRaMbLE). Growth characterization and genome sequencing showed that the ~358 kb fragment on Chr2 was accountable for the rapid growth of V. natriegens. The enabling tools we developed can help identify genomic regions that influence the rapid growth of V. natriegens without a prior understanding of genome mechanisms. This groundbreaking demonstration may also be extended to other organisms such as Escherichia coli, Pseudomonas putida, Bacillus subtilis, and so on.

Differential gene expression analysis reveals the fast-growth mechanisms in Melia dubia at different stand ages.

BACKGROUND: Melia dubia Cav. is a fast-growing multipurpose tree suitable for agroforestry and has been widely cultivated for wood-based industries, particularly pulp and paper production. Despite its high economic value in India, there is a lack of information regarding the molecular mechanism driving its fast-growth. Therefore, this study aimed to elucidate the molecular mechanisms responsible for fast-growth by expression analysis of selective candidate genes. METHODS AND RESULTS: Initially, growth traits were assessed, including tree height and diameter at breast height (DBH), across three different ages (one-year-old, two-year-old, and three-year-old) of M. dubia plantations. Tree volume based on tree height and DBH, was also calculated. The analysis of annual tree height increment revealed that the second-year plantation exhibited the higher increment, followed by first and third years. In contrast, DBH was maximum in third-year plantation, followed by the second and first years. Similarly, annual tree volume increment showed a similar trend with DBH that maximum in the third year, followed by second and first years. Furthermore, a differential gene expression analysis was performed using qRT-PCR on four genes such as Phloem Intercalated with Xylem (PXY), Clavata3/Embryo Surrounding Region-Related 41 (CLE41), 1-aminocyclopropane-1-carboxylic acid synthase (ACS-1) and Hemoglobin1 (Hb1) for downstream analysis. The relative gene expression showed up-regulation of CLE41, ACS-1, and Hb1 genes, while the PXY gene was downregulated across the tree ages. Interestingly, a positive association was observed between tree growth and the expression of the selected candidate genes. CONCLUSION: Our results pave the way for further research on the regulatory mechanisms of genes involved in fast-growth and provide a basis for genetic improvement of Melia dubia.

Effects of genetic selection for fast growth on the development of wooden breast myopathy in broilers.

1. This study investigated the physiological and molecular mechanisms leading to wooden breast (WB) by comparing growth parameters, oxygen consumption rate, thyroid hormone and gene expression patterns in fast- versus slow-growing broiler lines (Cobb500 and L1986, respectively).2. WB was observed in Cobb500 broilers only and was first diagnosed on d 21 post-hatch. Compared to the slow-growing L1986, Cobb500 showed a significantly higher growth rate, relative breast weight, breast thickness, meat pH and water-retention capacity (drip loss). Correspondingly, there was significantly lower relative heart weight, relative right ventricular weight, triiodothyronine and thyroxine concentrations and oxygen consumption rate.3. Compared to No-WB Cobb500, the WB-affected samples exhibited higher relative breast weight, breast thickness and drip loss and lower plasma total thyroxine (T4) concentrations.4. Selection for fast growth was associated with differential expression of genes involved in hypoxia (PLOD2), energy metabolism (FABP3, FABP4, CD36, and LPL), endoplasmic reticulum stress, muscle regeneration (CSRP3) and fibre-type switching (ANKRD1). WB-affected samples exhibited an upregulation of CSRP3, PLOD2 and ANKRD1, while CD36 was downregulated. Taken together, selection for fast growth and muscle gain is not matched by adequate cardiac and metabolic support systems.

Ultrafast Antisolvent Growth of Single-Crystal CsPbBr3 Microcavity for Whispering-Gallery-Mode Lasing.

In recent years, all-inorganic cesium lead bromide (CsPbBr3) perovskites have garnered considerable attention for their prospective applications in green photonics and optoelectronic devices. However, the development of efficient and economical methods to obtain high-quality micron-sized single-crystalline CsPbBr3 microplatelets (MPs) has become a challenge. Here, we report the synthesis of CsPbBr3 MPs on Si/SiO2 substrate by optimizing the ultrafast antisolvent method (FAS). This technique is able to produce well-dispersed, uniformly sized, and morphologically regular tetragonal phase single crystals, which can give strong green emission at room temperature, with excellent stability and excitonic character. Moreover, the crystals demonstrated lasing with a whispering gallery mode with a low threshold. These results suggest that the single-crystalline CsPbBr3 MPs synthesized by this method are of high optical quality, holding vast potential for future applications in photonic devices.

Identification and Characterization of PRE Genes in Moso Bamboo (Phyllostachys edulis).

Basic helix-loop-helix (bHLH)/HLH transcription factors are involved in various aspects of the growth and development of plants. Here, we identified four HLH genes, PePRE1-4, in moso bamboo plants that are homologous to Arabidopsis PRE genes. In bamboo seedlings, PePRE1/3 were found to be highly expressed in the internode and lamina joint by using quantitative RT-PCR analysis. In the elongating internode of bamboo shoots, PePRE genes are expressed at higher levels in the basal segment than in the mature top segment. Overexpression of PePREs (PePREs-OX) in Arabidopsis showed longer petioles and hypocotyls, as well as earlier flowering. PePRE1 overexpression restored the phenotype due to the deficiency of AtPRE genes caused by artificial micro-RNA. PePRE1-OX plants showed hypersensitivity to propiconazole treatment compared with the wild type. In addition, PePRE1/3 but not PePRE2/4 proteins accumulated as punctate structures in the cytosol, which was disrupted by the vesicle recycling inhibitor brefeldin A (BFA). PePRE genes have a positive function in the internode elongation of moso bamboo shoots, and overexpression of PePREs genes promotes flowering and growth in Arabidopsis. Our findings provided new insights about the fast-growing mechanism of bamboo shoots and the application of PRE genes from bamboo.

Ultrafast One-Step Deposition Route to Fabricate Single-Crystal CsPbX3 (X = Cl, Cl/Br, Br, and Br/I) Photodetectors.

Inorganic perovskites have received much attention due to their stability and high performance in luminescence, photoelectric conversion, and photodetection. However, perovskite optoelectronic devices prepared by the solution technique are still suffering from time-consuming and complex operations. In this paper, a single-crystal perovskite-based photodetector (PD) is prepared by very fast one-step deposition of synthesizing microplatelets (MPs) on the electrode directly. The saturated precursor is carefully optimized by adding appropriate antisolvent chlorobenzene (CB) to fabricate the MPs with their PL wavelength ranging from 418 to 600 nm. Furthermore, the PDs with a low dark current on order of nanoangstroms, high responsivity and detectivity of up to 10.7 A W-1 and 1012 Jones, respectively, and an ultrafast response rate featured by 278/287 µs (rise/decay time) are achieved. These all-inorganic perovskite PDs with a simple fabricating process and tunable detection wavelength meet the evolution tendency of PDs toward low cost and high performance, which is a high-profile strategy to realize high-performance perovskite PDs.

Methylomonas rapida sp. nov., a novel species of fast-growing, carotenoid-producing obligate methanotrophs with high biotechnological potential.

The genus Methylomonas accommodates strictly aerobic, obligate methanotrophs, with their sole carbon and energy sources restricted to methane and methanol. These bacteria inhabit oxic-anoxic interfaces of various freshwater habitats and have attracted considerable attention as potential producers of a single-cell protein. Here, we characterize two fast-growing representatives of this genus, strains 12 and MP1T, which are phylogenetically distinct from the currently described Methylomonas species (94.0-97.3 % 16S rRNA gene sequence similarity). Strains 12 and MP1T were isolated from freshwater sediments collected in Moscow and Krasnodar regions, respectively. Cells of these strains are Gram-negative, red-pigmented, highly motile thick rods that contain a type I intracytoplasmic membrane system and possess a particulate methane monooxygenase (pMMO) enzyme. These bacteria grow between 8 and 45 °C (optimum 35 °C) in a relatively narrow pH range of 5.5-7.3 (optimum pH 6.6-7.2). Major carotenoids synthesized by these methanotrophs are 4,4'-diaplycopene-4,4'-dioic acid, 1,1'-dihydroxy-3,4-didehydrolycopene and 4,4'-diaplycopenoic acid. High biomass yield, of up to 3.26 g CDW/l, is obtained during continuous cultivation of MP1T on natural gas in a bioreactor at a dilution rate of 0.22 h-1. The complete genome sequence of strain MP1T is 4.59 Mb in size; the DNA G + C content is 52.8 mol%. The genome encodes four rRNA operons, one pMMO operon and 4,216 proteins. The genome sequence displays 82-85 % average nucleotide identity to those of earlier described Methylomonas species. We propose to classify these bacteria as representing a novel species of the genus Methylomonas, M. rapida sp. nov., with the type strain MP1T (=KCTC 92586T = VKM B-3663T).

Rapid Growth of Metal-Metal Oxide Core-Shell Structures through Joule Resistive Heating: Morphological, Structural, and Luminescence Characterization.

The aim of this study is to prove that resistive heating enables the synthesis of metal/metal oxide composites in the form of core-shell structures. The thickness and morphology of the oxide layer depends strongly on the nature of the metal, but the influences of parameters such as the time and current profiles and the presence of an external field have also been investigated. The systems chosen for the present study are Zn/ZnO, Ti/TiO2, and Ni/NiO. The characterization of the samples was performed using techniques based on scanning electron microscopy (SEM). The thicknesses of the oxide layers varied from 10 µm (Zn/ZnO) to 50 µm (Ni/NiO). In the case of Zn- and Ti-based composites, the growth of nanostructures on the oxide layer was observed. Micro- and nanoneedles formed on the ZnO layer while prism-like structures appeared on the TiO2. In the case of the NiO layer, micro- and nanocrystals were observed. Applying an external electric field seemed to align the ZnO needles, whereas its effect on TiO2 and NiO was less appreciable, principally affecting the shape of their grain boundaries. The chemical compositions were analysed using X-ray spectroscopy (EDX), which confirmed the existence of an oxide layer. Structural information was obtained by means of X-ray diffraction (XRD) and was later checked using Raman spectroscopy. The oxide layers seemed to be crystalline and, although some non-stoichiometric phases appeared, the stoichiometric phases were predominant; these were wurtzite, rutile, and cubic for Zn, Ti, and Ni oxides, respectively. The photoluminescence technique was used to study the distribution of defects on the shell, and mainly visible bands (2-2.5 eV), attributed to oxygen vacancies, were present. The near-band edges of ZnO and TiO2 were also observed around 3.2-3.3 eV.

Transcriptomic signals of mitochondrial dysfunction and OXPHOS dynamics in fast-growth chicken.

Introduction: Birds are equipped with unique evolutionary adaptations to counter oxidative stress. Studies suggest that lifespan is inversely correlated with oxidative damage in birds. Mitochondrial function and performance are critical for cellular homeostasis, but the age-related patterns of mitochondrial gene expression and oxidative phosphorylation (OXPHOS) in birds are not fully understood. The domestic chicken is an excellent model to understand aging in birds; modern chickens are selected for rapid growth and high fecundity and oxidative stress is a recurring feature in chicken. Comparing fast- and slow-growing chicken phenotypes provides us an opportunity to disentangle the nexus of oxidative homeostasis, growth rate, and age in birds. Methods and Results: We compared pectoralis muscle gene expression patterns between a fast and a slow-growing chicken breed at 11 and 42 days old. Using RNAseq analyses, we found that mitochondrial dysfunction and reduced oxidative phosphorylation are major features of fast-growth breast muscle, compared to the slow-growing heritage breed. We found transcriptomic evidence of reduced OXPHOS performance in young fast-growth broilers, which declined further by 42 days. Discussion: OXPHOS performance declines are a common feature of aging. Sirtuin signaling and NRF2 dependent oxidative stress responses support the progression of oxidative damage in fast-growth chicken. Our gene expression datasets showed that fast growth in early life places immense stress on oxidative performance, and rapid growth overwhelms the OXPHOS system. In summary, our study suggests constraints on oxidative capacity to sustain fast growth at high metabolic rates, such as those exhibited by modern broilers.

Total and Mitochondrial Transcriptomic and Proteomic Insights into Regulation of Bioenergetic Processes for Shoot Fast-Growth Initiation in Moso Bamboo.

As a fast-growing, woody grass plant, Moso bamboo (Phyllostachys edulis) can supply edible shoots, building materials, fibrous raw material, raw materials for crafts and furniture and so on within a relatively short time. Rapid growth of Moso bamboo occurs after the young bamboo shoots are covered with a shell and emerge from the ground. However, the molecular reactions of bioenergetic processes essential for fast growth remain undefined. Herein, total and mitochondrial transcriptomes and proteomes were compared between spring and winter shoots. Numerous key genes and proteins responsible for energy metabolism were significantly upregulated in spring shoots, including those involved in starch and sucrose catabolism, glycolysis, the pentose phosphate pathway, the tricarboxylic acid cycle and oxidative phosphorylation. Accordingly, significant decreases in starch and soluble sugar, higher ATP content and higher rates of respiration and glycolysis were identified in spring shoots. Further, the upregulated genes and proteins related to mitochondrial fission significantly increased the number of mitochondria, indirectly promoting intracellular energy metabolism. Moreover, enhanced alternate-oxidase and uncoupled-protein pathways in winter shoots showed that an efficient energy-dissipating system was important for winter shoots to adapt to the low-temperature environment. Heterologous expression of PeAOX1b in Arabidopsis significantly affected seedling growth and enhanced cold-stress tolerance. Overall, this study highlights the power of comparing total and mitochondrial omics and integrating physiochemical data to understand how bamboo initiates fast growth through modulating bioenergetic processes.

Overexpression of DsEXLA2 gene from Dendrocalamus sinicus accelerates the plant growth rate of Arabidopsis.

Expansins play crucial roles in cell wall loosening and a range of life activities involving cell wall modification. Nevertheless, the biological functions of expansin genes during fast growth of bamboo remain unclear. In this study, Dendrocalamus sinicus, the largest and fastest growing bamboo species in the world, was used as the research material, and the full length of DsEXLA2 was cloned. Bioinformatics analysis revealed that DsEXLA2 contained expansin family typical domains (DPBB_1 and Pollen_allerg_1, CDRC motif) and amino acid sequence was highly conserved among different species. The expression level of DsEXLA2 increased from top section to basal section in different internodes. Subcellular localization verified that DsEXLA2 protein was located in the cell wall. Further genetic transformation studies in Arabidopsis indicated that compared with the wild type, DsEXLA2 overexpressed transgenic plants exhibited higher plant height, thicker stem, larger leaf, and less epidermal hair number and smaller stomatal aperture in the prophase and metaphase of growth. In addition, the cellulose content in the stem of transgenic plants was increased, and cell wall was thickened significantly. Moreover, a total of 1656 differentially expressed genes (DEGs) were identified by RNA-seq. The upregulated genes were predominantly enriched in the plant-pathogen interaction, MAPK signaling pathway-plant, plant hormone signal transduction, lipid metabolism and amino acid metabolism, while the downregulated genes were mainly enriched in energy metabolism, carbohydrate metabolism, plant hormone signal transduction and ribosome. These data implied that overexpression of DsEXLA2 gene accelerates the plant growth rate of Arabidopsis. This study is helpful to reveal the molecular mechanism of DsEXLA2 in culm growth and development of D. sinicus, and to understand the rapid growth of bamboos.

Engineering Natural Competence into the Fast-Growing Cyanobacterium Synechococcus elongatus Strain UTEX 2973.

Natural transformation is the process by which bacteria actively take up and integrate extracellular DNA into their genomes. In cyanobacteria, natural transformation has only been experimentally demonstrated in a few species. Although cyanobacteria are important model systems for studying photosynthesis and circadian cycling, natural transformation in cyanobacteria has not been characterized to the degree that the process has been studied in other Gram-negative bacteria. Two cyanobacterial species that are 99.8% genetically identical provide a unique opportunity to better understand the nuances of natural transformation in cyanobacteria: Synechococcus elongatus PCC 7942 and Synechococcus elongatus UTEX 2973 (hereafter called Synechococcus 7942 and Synechococcus 2973, respectively). Synechococcus 7942 is a naturally transformable model system, while Synechococcus 2973 is a recently discovered species that is not naturally competent. Taking only 1.5 h to replicate, Synechococcus 2973 is the fastest-growing cyanobacterial species known and thus is a strong candidate for serving as a model organism. However, its inability to undergo natural transformation has prevented it from becoming a widely used model system. By substituting polymorphic alleles from Synechococcus 7942 for native Synechococcus 2973 alleles, natural transformation was introduced into Synechococcus 2973. Two genetic loci were found to be involved in differential natural competence between the two organisms: transformation pilus component pilN and circadian transcriptional master regulator rpaA. By using targeted genome editing and enrichment outgrowth, a strain that was both naturally transformable and fast-growing was created. This new Synechococcus 2973-T strain will serve as a valuable resource to the cyanobacterial research community. IMPORTANCE Certain bacterial species have the ability to take up naked extracellular DNA and integrate it into their genomes. This process is known as natural transformation and is widely considered to play a major role in bacterial evolution. Because of the ease of introducing new genes into naturally transformable organisms, this capacity is also highly valued in the laboratory. Cyanobacteria are photosynthetic and can therefore serve as model systems for some important aspects of plant physiology. Here, we describe the creation of a modified cyanobacterial strain (Synechococcus 2973-T) that is capable of undergoing natural transformation and has a replication time on par with that of the fastest-growing cyanobacterium discovered to date. This new cyanobacterium has the potential to serve as a new model organism for the cyanobacterial research community and will allow experiments to be completed in a fraction of the time it has taken to complete previous assays.

Molecular characterization of fast-growing melanomas.

BACKGROUND: The rate of growth of primary melanoma is a robust predictor of aggressiveness, but the mutational profile of fast-growing melanomas (FGMM) and the potential to stratify patients at high risk of death has not been comprehensively studied. OBJECTIVE: To investigate the epidemiologic, clinical, and mutational profile of primary cutaneous melanomas with a thickness ≥ 1 mm, stratified by rate of growth. METHODS: Observational prospective study. Deep-targeted sequencing of 40 melanoma driver genes on formalin fixed, paraffin-embedded primary melanoma samples. Comparison of FGMM (rate of growth > 0.5 mm/month) and nonFGMM (rate of growth ≤ 0.5 mm/month). RESULTS: Two hundred patients were enrolled, among wom 70 had FGMM. The relapse-free survival was lower in the FGMM group (P = .014). FGMM had a higher number of predicted deleterious mutations within the 40 genes than nonFGMM (P = .033). Ulceration (P = .032), thickness (P = .006), lower sun exposure (P = .049), and fibroblast growth factor receptor 2 (FGFR2) mutations (P = .037) were significantly associated with fast growth. LIMITATIONS: Single-center study, cohort size, potential memory bias, number of investigated genes. CONCLUSION: Fast growth is linked to specific tumor biology and environmental factors. Ulceration, thickness, and FGFR2 mutations are associated with fast growth. Screening for FGFR2 mutations might provide an additional tool to better identify FGMM, which are probably good candidates for adjuvant therapies.

Genome Structural Variation Landscape and Its Selection Signatures in the Fast-growing Strains of the Pacific Oyster, Crassostrea gigas.

The Pacific oyster (Crassostrea gigas) genome is highly polymorphic and affluent in structural variations (SVs), a significant source of genetic variation underlying inter-individual differences. Here, we used two genome assemblies and 535 individuals of genome re-sequencing data to construct a comprehensive landscape of structural variations in the Pacific oyster. Through whole-genome alignment, 11,087 short SVs and 11,561 copy number variations (CNVs) were identified. While analysis of re-sequencing data revealed 511,170 short SVs and 979,486 CNVs, a total of 63,100 short SVs and 58,182 CNVs were identified in at least 20 samples and regarded as common variations. Based on the common short SVs, both Fst and Pi ratio statistical methods were employed to detect the selective sweeps between 20 oyster individuals from the fast-growing strain and 20 individuals from their corresponding wild population. A total of 514 overlapped regions (8.76 Mb), containing 746 candidate genes, were identified by both approaches, in addition with 103 genes within 61 common CNVs only detected in the fast-growing strains. The GO enrichment and KEGG pathway analysis indicated that the identified candidate genes were mostly associated with apical part of cell and were significantly enriched in several metabolism-related pathways, including tryptophan metabolism and histidine metabolism. This work provided a comprehensive landscape of SVs and revealed their responses to selection, which will be valuable for further investigations on genome evolution under selection in the oysters.

Integrative lncRNA landscape reveals lncRNA-coding gene networks in the secondary cell wall biosynthesis pathway of moso bamboo (Phyllostachys edulis).

BACKGROUND: LncRNAs are extensively involved in plant biological processes. However, the lack of a comprehensive lncRNA landscape in moso bamboo has hindered the molecular study of lncRNAs. Moreover, the role of lncRNAs in secondary cell wall (SCW) biosynthesis of moso bamboo is elusive. RESULTS: For comprehensively identifying lncRNA throughout moso bamboo genome, we collected 231 RNA-Seq datasets, 1 Iso-Seq dataset, and 1 full-length cDNA dataset. We used a machine learning approach to improve the pipeline of lncRNA identification and functional annotation based on previous studies and identified 37,009 lncRNAs in moso bamboo. Then, we established a network of potential lncRNA-coding gene for SCW biosynthesis and identified SCW-related lncRNAs. We also proposed that a mechanism exists in bamboo to direct phenylpropanoid intermediates to lignin or flavonoids biosynthesis through the PAL/4CL/C4H genes. In addition, we identified 4 flavonoids and 1 lignin-preferred genes in the PAL/4CL/C4H gene families, which gained implications in molecular breeding. CONCLUSIONS: We provided a comprehensive landscape of lncRNAs in moso bamboo. Through analyses, we identified SCW-related lncRNAs and improved our understanding of lignin and flavonoids biosynthesis.

Comparative Genomic and Transcriptomic Analysis Reveals Specific Features of Gene Regulation in Kluyveromyces marxianus.

Kluyveromyces marxianus is a promising host for producing bioethanol and heterologous proteins. It displays many superior traits to a conventional industrial yeast species, Saccharomyces cerevisiae, including fast growth, thermotolerance and the capacity to assimilate a wider variety of sugars. However, little is known about the mechanisms underlying the fast-growing feature of K. marxianus. In this study, we performed a comparative genomic analysis between K. marxianus and other Saccharomycetaceae species. Genes involved in flocculation, iron transport, and biotin biosynthesis have particularly high copies in K. marxianus. In addition, 60 K. marxianus specific genes were identified, 45% of which were upregulated during cultivation in rich medium and these genes may participate in glucose transport and mitochondrion related functions. Furthermore, the transcriptomic analysis revealed that under aerobic condition, normalized levels of genes participating in TCA cycles, respiration chain and ATP biosynthesis in the lag phase were higher in K. marxianus than those in S. cerevisiae. Levels of highly copied genes, genes involved in the respiratory chain and mitochondrion assembly, were upregulated in K. marxianus, but not in S. cerevisiae, in later time points during cultivation compared with those in the lag phase. Notably, during the fast-growing phase, genes involved in the respiratory chain, ATP synthesis and glucose transport were co-upregulated in K. marxianus. A few shared motifs in upstream sequences of relevant genes might result in the co-upregulation. Specific features in the co-regulations of gene expressions might contribute to the fast-growing phenotype of K. marxianus. Our study underscores the importance of genome-wide rewiring of the transcriptional network during evolution.

Developing rapid growing Bacillus subtilis for improved biochemical and recombinant protein production.

Bacillus subtilis is a model Gram-positive bacterium, which has been widely used as industrially important chassis in synthetic biology and metabolic engineering. Rapid growth of chassis is beneficial for shortening the fermentation period and enhancing production of target product. However, engineered B. subtilis with faster growth phenotype is lacking. Here, fast-growing B. subtilis were constructed through rational gene knockout and adaptive laboratory evolution using wild type strain B. subtilis 168 (BS168) as starting strain. Specifically, strains BS01, BS02, and BS03 were obtained through gene knockout of oppD, hag, and flgD genes, respectively, resulting 15.37%, 24.18% and 36.46% increases of specific growth rate compared with BS168. Next, strains A28 and A40 were obtained through adaptive laboratory evolution, whose specific growth rates increased by 39.88% and 43.53% compared to BS168, respectively. Then these two methods were combined via deleting oppD, hag, and flgD genes respectively on the basis of evolved strain A40, yielding strain A4003 with further 7.76% increase of specific growth rate, reaching 0.75 h-1 in chemical defined M9 medium. Finally, bioproduction efficiency of intracellular product (ribonucleic acid, RNA), extracellular product (acetoin), and recombinant proteins (green fluorescent protein (GFP) and ovalbumin) by fast-growing strain A4003 was tested. And the production of RNA, acetoin, GFP, and ovalbumin increased 38.09%, 5.40%, 9.47% and 19.79% using fast-growing strain A4003 as chassis compared with BS168, respectively. The developed fast-growing B. subtilis strains and strategies used for developing these strains should be useful for improving bioproduction efficiency and constructing other industrially important bacterium with faster growth phenotype.

Biochemical Characteristics and a Genome-Scale Metabolic Model of an Indian Euryhaline Cyanobacterium with High Polyglucan Content.

Marine cyanobacteria are promising microbes to capture and convert atmospheric CO2 and light into biomass and valuable industrial bio-products. Yet, reports on metabolic characteristics of non-model cyanobacteria are scarce. In this report, we show that an Indian euryhaline Synechococcus sp. BDU 130192 has biomass accumulation comparable to a model marine cyanobacterium and contains approximately double the amount of total carbohydrates, but significantly lower protein levels compared to Synechococcus sp. PCC 7002 cells. Based on its annotated chromosomal genome sequence, we present a genome scale metabolic model (GSMM) of this cyanobacterium, which we have named as iSyn706. The model includes 706 genes, 908 reactions, and 900 metabolites. The difference in the flux balance analysis (FBA) predicted flux distributions between Synechococcus sp. PCC 7002 and Synechococcus sp. BDU130192 strains mimicked the differences in their biomass compositions. Model-predicted oxygen evolution rate for Synechococcus sp. BDU130192 was found to be close to the experimentally-measured value. The model was analyzed to determine the potential of the strain for the production of various industrially-useful products without affecting growth significantly. This model will be helpful to researchers interested in understanding the metabolism as well as to design metabolic engineering strategies for the production of industrially-relevant compounds.

Amino acids are key substrates to Escherichia coli BW25113 for achieving high specific growth rate.

Studying substrate consumption in nutrient-rich conditions is challenging because often the growth medium includes undefined components like yeast extract or peptone. For clear and consistent results, it is necessary to use defined medium, where substrate utilization can be followed. In the present work, Escherichia coli BW25113 batch growth in a medium supplemented with 20 proteinogenic amino acids and glucose was studied. Focus was on the quantitative differences in substrate consumption and proteome composition between minimal and nutrient-rich medium. In the latter, 72% of carbon used for biomass growth came from amino acids and 28% from glucose. Serine was identified as the most consumed substrate with 41% of total carbon consumption. Proteome comparison between nutrient-rich and minimal medium revealed changes in TCA cycle and acetate producing enzymes that together with extracellular metabolite data pointed to serine being consumed mainly for energy generation purposes. Serine removal from the growth medium decreased specific growth rate by 22%. In addition, proteome comparison between media revealed a large shift in amino acid synthesis and translation related proteins. Overall, this work describes in quantitative terms the batch growth carbon uptake profile and proteome allocation of E. coli BW25113 in minimal and nutrient-rich medium.

A commentary on the impacts of 'Great Lockdown' and its aftermath on scaling firms: What are the implications for entrepreneurial research?

This commentary argues that scaling fast growth firms drive economic development, even in recessionary periods. While the coronavirus induced the 'Great Lockdown' and its aftermath poses particular challenges, we argue that the crisis presents the entrepreneurial scholarly community with an opportunity to re-orientate our research. Rather than more narratives of business success in the face of adversity, the Great Lockdown presents us with a fresh opportunity to examine how scaling is affected by context, by luck and by the porous nature of business growth. In so doing, our hope is that it will encourage our community to adopt a more proactive agenda to support policy makers and entrepreneurs.