Biotechnology Notes最新文献

Insect olfaction directly impacts insect behavior and thus is an important consideration in the development of smart farming tools and in integrated pest management strategies. Insect olfactory receptors (ORs) have been traditionally studied using Drosophila empty neuron systems or with expression and functionalization in HEK293 cells or Xenopus laevis oocytes. Recently, the yeast Saccharomyces cerevisiae (S. cerevisiae) has emerged as a promising chassis for the functional expression of heterologous seven transmembrane receptors. S. cerevisiae provides a platform for the cheap and high throughput study of these receptors and potential deorphanization. In this study, we explore the foundations of a scalable yeast-based platform for the functional expression of insect olfactory receptors by employing a genetically encoded calcium sensor for quantitative evaluation of fluorescence and optimized experimental parameters for enhanced functionality. While the co-receptor of insect olfactory receptors remains non-functional in our yeast-based system, we thoroughly evaluated various experimental variables and identified future research directions for establishing an OR platform in S. cerevisiae.

SynBio Africa is a forum for researchers, students, citizen scientists, policymakers and the public to convene and develop successful pathways for the propagation of synthetic biology technologies, products, and services throughout Africa. Our vision is to have a healthy, safe, and sustainable world through synthetic biology. In Africa, synthetic biology has the potential to greatly contribute to national development agenda through the following ways: i) by anchoring a sustainable bioeconomy; ii) by helping develop innovative medicines; iii) by reducing pollution, and iv) by increasing crop production to reduce hunger. However, there is little to no information on synthetic biology and its regulatory policies in Africa. Across the continent, scientists, policy makers, researchers and others are still working in silos—only partaking in consultative meetings to try and develop a set of unified policy guidelines. SynBio Africa is therefore proposing to establish the first Center of Excellence in Synthetic Biology in Africa with six themes, namely: research, capacity development, innovation hub, biosafety and biosecurity, and bioinformatics and data science, and one-health. Accordingly, SynBio Africa will work with collaborators from government and non-governmental organizations, the public and private sectors, and educational institutions from Uganda, Africa, and around the world to implement these six themes.

Yarrowia lipolytica is a modern workhorse for biotechnology that is amenable to genetic manipulations and can produce high levels of various enzymes. The present study was designed to engineer Y. lipolytica for the overexpression of α-bisabolene, a valuable biofuel precursor and pharmaceutical, making use of this yeast's ability to accumulate lipids, and with the use of a golden gate DNA assembly (GG) toolbox. By transforming Y. lipolytica with a GG genetic construct involving truncated 3-hydroxy-3-methyl-glutaryle coenzyme A reductase (tHMG) and α-bisabolene synthase (Bis) genes controlled by the strong TEF promoter and Lip2 terminator, the engineered yeast was able to produce 489 mg l⁻¹ of α-bisabolene. This was increased to 816 mg l⁻¹ by transforming a lipid-over-accumulating Y. lipolytica strain with the same genetic construct. Higher production titers of up to 1243 mg l⁻¹ could be also achieved by varying the culture conditions of the transformed strains.

Fe doped MgO nanoparticles were synthesized using a straightforward soft chemical method. We conducted a comprehensive examination of the electrical properties of Fe-doped MgO nanoparticles with a crystalline size range of 7–10 nm. Simultaneously, we explored their antibacterial capabilities. Our findings indicate that an increase in the concentration of Fe-doped MgO correlates with an enhanced bactericidal effect. To gain a deeper understanding of charge transfer processes, the AC conductivity and dielectric characteristics of the samples across various temperatures and frequencies was studied.The antibacterial activity was studied utilising the MIC methodology, the live count (LC) method, and the agar cup technique in addition to the electrical characteristics. After exposure to nanoparticles, we observed the disruption of pathogenic cell walls through transmission electron microscopy (TEM) analysis. These results suggest that Fe-doped MgO nanoparticles hold promise for the development of novel, more effective antibacterial drugs. The ½ MIC for E.coli was found to be 2.75 μg/ml, while for Bacillus sp., it was 1.75 μg/ml when exposed to Fe-doped MgO nanoparticles. This dosage level may find applications in the medical field. However, further investigations are required to assess potential toxicity and long-term environmental and human health effects. If successful in vivo tests follow, Fe-doped MgO nanoparticles could emerge as valuable antibacterial agents.

Enhanced synthesis of hyaluronic acid (HA) with recombinant Corynebacterium glutamicum as production host was achieved in this work. Hyaluronan synthase (HAS), which is a membrane protein acting as a key enzyme in HA biosynthesis, impacts both HA yield and its molecular weight. Cell morphology, which includes size, shape, and surface area, has a large impact on the expression and activity of HAS. Therefore, deliberate regulation of cell morphology holds the potential to enhance HA production. Here, we constructed three modules, namely the transporter module, the morphology tuning module and the HA synthesis module. The transporter module contains a strong constitutive promoter P_tuf and arabinose transport protein was used to control the maximum amount of inducer entering the cell, thus reducing excessive cell deformation. The morphology tuning module contains an arabinose-inducible weak promoter P_BAD and a cell-division-relevant gene was used to sense intracellular inducer concentrations and achieve different degrees of change in cell size. These two modules worked together, described as a dual-valve regulation, to achieve fine-tuning of cell morphology, resulting in a 1.87-fold increase in cell length and a 2.08-fold increase in cell membrane. When combined with the HA synthesis module, the HA titer reached 16.0 g/L, which was 1.6 times the yield reported in the previous morphology-engineered strain. Hence, for the first time, a morphologically engineered strain resulting in both high cell density and HA titer was constructed.

Circular mRNA (circmRNA) is a covalent closed loop formed by reverse splicing of the 3′ end to the 5′ end of mRNA. Compared to traditional linear mRNAs, circmRNAs can mediate efficient, stable, and durable protein expression and are considered an alternative to linear mRNAs in terms of therapeutic reagents. With the continuous development of circmRNA research, circmRNA has also made significant progress in vaccines and cellular therapies. In this review, we present research advances in the in vitro synthesis of circmRNAs, focusing on the biological ligation methods of circmRNAs and current applications, with a summary of challenges regarding circmRNA design, synthesis, and applications. Based on the enhanced stability of circmRNAs, further research on circmRNAs could help expand their applications in biotherapeutics and strengthen their role in basic medical applications.

Dengue fever is caused by any of the four serotypes of dengue viruses, DENV-1, DENV-2, DENV-3 and DENV-4 spread by mosquito bites and is important to distinguish between them due to lack of cross-protective neutralizing antibodies for each serotype. Secondary infections also put individuals at higher risk for severe dengue illness than those who have not been previously infected. Current preferred assays include reverse transcription-PCR (RT-PCR) and ELISA. To enable on-field diagnosis of dengue serotypes, the detection process would need to be simplified or at least semi-automated. A downstream detection module was conceptualized and fabricated to detect the amplified DNA from the provided PCR mix (product) of previously developed modular microfluidic chips involving sample loading, cell lysis, RNA extraction and RT-PCR. Further, to ensure accuracy, each serotype assay necessitates a positive control, negative control and test sample, which constitutes 3 separate channels for the diagnosis of just 1 serotype. In this study, a 6-channel bi-assay microfluidic chip was designed with pre-loaded diluent and cyanine dye, sample chamber for loading, sequential fluidic sample mixing, and integrated membranes for simultaneous (6-channel) fluidic manipulation from a single actuation source. Positive samples will turn the dye from blue to violet while the negative controls will remain blue. The integrated membranes provided color contrast and facilitated the manipulation of the samples to the same line of sight for simultaneous analysis, paving the way for automated color analysis via smartphone.

Synthetic biology is an engineering discipline that applies engineering principles to rationally design novel biological systems. It has the potential to contribute to solving major global challenges in a multitude of areas, from healthcare to sustainability. While the engineering biology landscape is robust and well-established in certain countries, the ecosystem and infrastructure for genetic engineering in other countries, including Slovakia, are underdeveloped. Consequently, such countries are missing the major economic and social benefits that the practical applications of the rational design of biological systems may provide. In this work, we briefly assess the status of the synthetic biology landscape in Slovakia in different areas, including research efforts, industrial participation, governmental policy, and the educational landscape. We describe the major challenges that the Slovak synthetic biology sector faces and propose a strategy that academics, policymakers, and industry could take to activate the proliferation of the Slovak synthetic biology ecosystem.

To improve the production of 3-HP with glucose as a substrate, the malonyl-CoA and propionyl-CoA pathways were coupled to regulate NADP⁺/NADPH regeneration in the recombinant E. coli. The strain Ec-AM that overexpressed the key enzymes of the malonyl-CoA pathway, acetyl CoA carboxylase (ACC) from Ustilago maydis and malonyl CoA reductase (MCR) from Chloroflexus aurantiacus, produced 0.26 g/L of 3-HP in 25-h shake flask cultivation. The strain Ec-P overexpressing the key enzyme of the propionyl-CoA pathway, propionyl-CoA dehydrogenase (PACD) from Candida rugosa, produced 0.11 g/L of 3-HP. However, 3-HP titer of the strain Ec-PAM overexpressing PACD along with ACC and MCR, via two pathways cooperation, was 1.29 g/L. The addition of biotin and bicarbonate improved the 3-HP production of the strain Ec-PAM. 3-HP titer of strain Ec-ΔY-ΔP-PAM with double deletion of ygfH (encoding propionyl-CoA: succinate-CoA transferase) and prpC (encoding methylcitrate synthase) genes reached 1.94 g/L, which was 1.5-fold higher than that of the strain Ec-PAM cultured under the same conditions.

Lipopeptides, novel biosurfactants showing versatile promising applications in enhanced oil recovery, textile industry, agriculture and daily chemical products, etc., are profoundly highlighted recently. Surfactin is one of the most typical representatives of lipopetide family. The critical micelle concentration (CMC) of surfactin is as low as 10–20 mg/L. When its concentration reaches above the CMC, different micelle structure will be formed and the surface-active performances might be changed with varied micelle morphologies. Thus, observation of the changes of surfactin micellar form at different concentrations is of great significance for its new applications. But so far, the micelle structure of surfactin (and also other lipopeptide molecules) is not reported yet, and the method for effectively observing the micelle morphology is limited as well. Here, we developed a method based on transmission electron microscopy combined with negative staining to observe the morphology of surfactin micelles, with which we can clearly observe the changes of micelle morphology of surfactin (or other lipopeptides) at different concentrations. Spherical micelles only form when the concentration of surfactin is low. With the increase in concentration, rod-shaped micelles of surfactin can form. Furthermore, complex rod-shaped-micelle-layer and big ring structure will form when the concentration of surfactin is very high.