Biotechnology Notes最新文献

Despite its start in the early 2000s, synthetic biology is still overall perceived as a young discipline. In some countries, such as the US, synthetic biology is academically and industrially established, while in others, including Germany, it is still an upcoming field of research. Issues with funding schemes, commercial translation of technologies, public perception, and regulations need to be addressed to establish synthetic biology as a key discipline of the 21^st century. This perspective article reviews the German and European synthetic biology landscape and how the German Association for Synthetic Biology (GASB) is addressing the above-mentioned challenges with its events and community-building activities.

Synthetic biology (SynBio) is a rapidly growing scientific discipline. In the Netherlands, various universities and companies are tackling a variety of opportunities and challenges within this field. In this perspective article, we review the current synthetic biology landscape in the Netherlands across academia, industry, politics, and society. Especially within Dutch academia there is an active, though only partially connected, research community involved in various domains of SynBio. Mostly supported by governmental funding, academic research is focusing on top-down synthetic biology, involving the engineering of, for example, bacteria and yeast for bioproduction, as well as bottom-up and cell-free synthetic biology aiming to understand life and build synthetic cells. There is also a large number of talented and motivated students interested in the field, exemplified by the participation and success of Dutch teams in the international iGEM synthetic biology competition. Commercial synthetic biology activities are taking place in various large industrial companies, as well as in start-ups and spin-offs, mostly divided over several ‘SynBio hubs’ in the Netherlands. However, the investment, regulatory and public-perception landscape is not yet optimal to stimulate entrepreneurial activities in SynBio. The Dutch and global society can further benefit from the large promise of SynBio through better integration of people active in the Dutch SynBio field, frequent political and public dialogue, and more attention towards regulatory issues. The recently founded Dutch synthetic biology association SynBioNL aims to contribute to realizing a positive impact on society by stimulating advances of the field in the Netherlands and beyond.

Dark fermentation is considered as one of the most practical biological hydrogen production methods. However, current productivity and yield are still not economically viable for industrial applications. This biological process must be improved through multiple strategies, of which screening for more effective microbial strains is an important aspect. Here, based on the hydrogen production pathway of E. aerogenes, we describe three strategies to improve hydrogen production by effectively regulating the anaerobic metabolism of E. aerogenes through genetic modification. This protocol describes in detail how to obtain NADH dehydrogenase-damaged mutants and overexpress Nad synthase genes using the CRISPR-Cas9 gene editing system. In addition, the overexpression of small RNA RyhB was achieved and verified by Northern Blot. This protocol is of great significance for the study of genetic engineering operation in E. aerogenes and other bacteria, and also provides theoretical guidance and technical support for the study of E. aerogenes biological hydrogen production.

Synthetic genomics, or engineering biology at the level of whole genomes and whole organisms, is an emerging outgrowth of parts-based synthetic biology. This nascent subfield is also diverse and difficult to characterize. As social scientists investigating responsible research and innovation in synthetic genomics, we suggest that focusing on the organism is a fruitful approach to making sense of the diversity it encompasses. Here, we offer a heuristic in the form of a tagging system to organize projects by the roles the engineered organism is asked to perform. We suggest several reasons why this system is useful for understanding the current shape and future directions of the field, especially in light of the need to ask: how does engineering biology contribute to building a future of sustainable relationships with other creatures?

Virus-like particles (VLPs) have a great application prospect in vaccines and molecule delivery carriers. In this study, in order to solve the problem of low expression and low assembly efficiency of VLPs, the conditions for the assembly and purification of eight representative VLPs (hepatitis B virus core antigen protein particles, Qbeta phage, MS2 phage, P22 phage, cowpea chlorotic mottle virus, tobacco Mosaic virus, ferritin and encapsulin) expressed in Escherichia coli were optimized. The VLPs with high expression, easy assembly and good purification properties were selected as the preferred objects for potential biological applications.

This study reports the development of a polymer-based catheter coating to facilitate controlled release of antimicrobial peptides (AMP) to target both planktonic bacteria and biofilm in the urinary catheter environment. Catheter associated urinary tract infection (CAUTI) is a common nosocomial infection among hospitalized patients and is a major reservoir of antimicrobial resistant pathogens. Although silver- or antibiotics-coated catheters have been deployed to minimise CAUTI, the inconsistency and lack of durability in antibacterial properties of these coatings have limited their clinical use. The incorporation of AMPs in catheter coatings has gained interest due to the effective bacteria killing effects of AMPs, with few reports on bacterial resistance development against peptides. This study aims to deploy a novel and potentially cost-effective technique to coat an anhydrous polymeric coating impregnated with AMPs for silicone-based catheters, to overcome limitations in conventional hydrogel-based coatings. Sustained peptide release was achieved with the development of an Ethyl Cellulose (EC): 1-Palmitoyl-2-oleoylphosphatidylcholine (POPC)-based diffusion layer over an AMP-laden Polycaprolactone (PCL)-based layer to control AMP diffusion into the environment over a clinically relevant duration. The ‘AMP-EC-PCL’ coating showed good anti-bacteria performance against uropathogenic Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa for up to 6 days. The coating also showed excellent anti-biofilm capability against green fluorescent protein (GFP)-tagged UTI E. coli. Fifteen centimeter catheter segments of single layer ‘AMP-EC-PCL’-coated catheters showed sustainable AMP release kinetics up to 7 days, where good antibacterial and anti-biofilm activity against E. coli was observed. The full scale ‘AMP-EC-PCL’-coated catheter showed improved mechanical integrity compared to commercial silicone catheters with preservation of the catheter balloon integrity upon expansion. Wound healing studies of the coated PDMS samples in mice models showed a reduction in bacteria concentration as compared to uncoated PDMS, indicating in vivo efficacy potential of the developed catheter coating platform.

Methylxanthines, including caffeine and theophylline, are a class of natural and synthetic compounds with important roles in food, cosmetics, and medicine. These compounds are metabolized by bacteria using five enzymes from the Rieske non-heme iron oxygenase family, NdmABCDE. The NdmCDE complex is responsible for the N₇-demethylation of 7-methylxanthine to xanthine and was originally described as being highly specific for 7-methylxanthine. Here, we report that the NdmCDE complex is also active toward theobromine, producing 3-methylxanthine due to N₇-demethylation. Minimal activity was observed when the enzyme complex was tested with caffeine or paraxanthine, indicating that the presence of the N₁-methyl group significantly inhibits N₇-demethylase activity by NdmCDE. We also demonstrated positional promiscuity in the N₃-demethylase, NdmB, which is able to carry out N₁-demethylation of paraxanthine. The N₁-demethylation by NdmB is limited to paraxanthine and was not observed when caffeine or theophylline were assayed. These newly discovered activities were observed when enzymes were overexpressed in E. coli and differ from results with purified enzymes assayed in vitro, indicating that they may behave differently in vivo. Furthermore, these results reveal promiscuity of bacterial N-demethylase enzymes that can be used to engineer new enzymes and bacterial strains for production of high-value methylxanthines.

Triterpenoids are a group of natural products with promising biological activities. Studying the transport of triterpenoids is of vital importance for designing novel microbial cell factory for efficient bioproduction of triterpenoids, as well as understanding the host defense mechanism. In this review, the distribution of triterpenoids was introduced at first. Then, the transport of triterpenoid, including the intracellular transport, transport across the cell membrane, and transport in host tissues, was thoroughly summarized. Finally, the challenges for the research in triterpenoid transport and possible solutions were also discussed.

Butyrate is a key microbial metabolite known to enhance host metabolic processes by reducing blood glucose levels and promoting energy metabolism, thereby potentially suppressing the risk of developing metabolic syndrome. In this study, we examined the activity of butyrate on modulating the endocannabinoid system, which regulates food intake and energy metabolism. Furthermore, we genetically engineered probiotics to produce butyrate to investigate their effects on the endocannabinoid system. Our study shows that the engineered probiotics exerted antagonistic effects on the endocannabinoid system by downregulating and upregulating the endocannabinoid-synthesizing and -degrading enzyme expression, respectively. Our results suggest that butyrate can modulate the endocannabinoid system, and incorporation of butyrate-producing bacteria into the gut microbiota can potentially aid in re-establishing homeostatic metabolic processes and alleviating metabolic syndrome.

Pearl millet [Pennisetum glaucum (L.) R. Br.] is a climate-hardy nutricereal and an essential staple for the people living in dry regions. Substantial improvement has been achieved for seed yield stability in pearl millet, the cultivable area under pearl millet reduces. Deployment of early-flowering, bold-seeded and dwarf genotypes in pearl millet is a vital breeding strategy to improve grain production and enhance the adaptability of pearl millet in low-input farms. Therefore, an experiment was performed for mapping agronomically important traits like flowering time (FT), plant height (PH), panicle length (PL), and 1000-grain weight (TGW) in 317 recombinant inbred line (RIL) population derived from ICMS 8511-S1-17-2-1-1-B-P03 × AIMP 92901-S1-183-2-2-B-08 cross. Broad-sense heritability estimates were high to very high, ranging from 0.52 (PL) to 0.86 (PH). FT showed a significant positive correlation with PH. A key QTL for FT was mapped on LG 1, 15 QTLs for PH scattered on 10 chromosomes, five QTLs for PL dispersed on four chromosomes, and two QTLs for TGW spanned linkage groups 3 and 7. One QTL on LG1 was common for FT and PH. Two major QTLs for PH, one each on LG4B/8 cM and LG7/110 cM were detected. The large effect QTL for TGW on LG7 had a phenotypic variance (R²) of 24.3%. The R² for PH and PL ranged between 5.2 - 24.5% and 5.0–11.5%, respectively. The QTLs mapped for FT and other agronomic traits in the current study can be exploited to develop elite hybrid parental genotypes/cultivars through marker-assisted breeding and genomic selection.