scholarly journals Biotechnology Applications of Cell-Free Expression Systems

Life ◽  
2021 ◽  
Vol 11 (12) ◽  
pp. 1367
Author(s):  
August Brookwell ◽  
Javin P. Oza ◽  
Filippo Caschera
Keyword(s):  
Low Cost ◽  
Free Expression ◽  
Expression Systems ◽  
Genetic Circuits ◽  
Growth Cycles ◽  
Platform Technology ◽  
Near Future ◽  
Cell Free Protein Synthesis

Cell-free systems are a rapidly expanding platform technology with an important role in the engineering of biological systems. The key advantages that drive their broad adoption are increased efficiency, versatility, and low cost compared to in vivo systems. Traditionally, in vivo platforms have been used to synthesize novel and industrially relevant proteins and serve as a testbed for prototyping numerous biotechnologies such as genetic circuits and biosensors. Although in vivo platforms currently have many applications within biotechnology, they are hindered by time-constraining growth cycles, homeostatic considerations, and limited adaptability in production. Conversely, cell-free platforms are not hindered by constraints for supporting life and are therefore highly adaptable to a broad range of production and testing schemes. The advantages of cell-free platforms are being leveraged more commonly by the biotechnology community, and cell-free applications are expected to grow exponentially in the next decade. In this study, new and emerging applications of cell-free platforms, with a specific focus on cell-free protein synthesis (CFPS), will be examined. The current and near-future role of CFPS within metabolic engineering, prototyping, and biomanufacturing will be investigated as well as how the integration of machine learning is beneficial to these applications.

The double and triple role of L-(+)-tartaric acid and dimethyl urea: A prevailing green approach in organic synthesis

2021 ◽  
Vol 25 ◽  
Author(s):  
Rashid Ali ◽  
Ajay Kumar Chinnam ◽  
Vikas R. Aswar
Keyword(s):  
Tartaric Acid ◽  
Organic Synthesis ◽  
Low Cost ◽  
Organic Transformations ◽  
Green Approach ◽  
Dimethyl Urea ◽  
Low Toxicity ◽  
The One ◽  
Near Future

: The deep eutectic mixtures (DESs), introduced as novel alternative to usual volatile organic solvents for organic transformations has attracted a tremendous attention of the research community because of their low cost, negligible vapour pressure, low toxicity, biodegradability, recyclability, insensitive towards moisture, and readily availability from bulk renewable resources. Although, the low melting mixture of dimethyl urea (DMU)/L-(+)-tartaric acid (TA) is still infancy yet much effective as it play double and triple roles such as solvent, catalyst and/or reagent in a same pot for many crucial organic transformations. These unique properties of DMU/TA mixture prompted us to provide a quick overview of where the field stands presently, and where it might be going in near future. To our best knowledge, no review dealing with the applications of a low melting mixture of DMU/TA appeared in the literature except the one published in 2017 describing only the chemistry of indole systems. Therefore, we intended to reveal the developments of this versatile low melting mixture in the modern organic synthesis since its first report in 2011 by Köenig’s team to till date. Hopefully, the present review article will be useful to the researcher working not only in the arena of synthetic organic chemistry but also to the scientists working in other branches of science and technology.

scholarly journals Locus Coeruleus Modulates Neuroinflammation in Parkinsonism and Dementia

2020 ◽  
Vol 21 (22) ◽  
pp. 8630
Author(s):  
Filippo Sean Giorgi ◽  
Francesca Biagioni ◽  
Alessandro Galgani ◽  
Nicola Pavese ◽  
Gloria Lazzeri ◽  
...  
Keyword(s):  
Experimental Data ◽  
Locus Coeruleus ◽  
Clinical Symptoms ◽  
Experimental Studies ◽  
Anatomy And Physiology ◽  
The Central Nervous System ◽  
Near Future

Locus Coeruleus (LC) is the main noradrenergic nucleus of the central nervous system, and its neurons widely innervate the whole brain. LC is severely degenerated both in Alzheimer’s disease (AD) and in Parkinson’s disease (PD), years before the onset of clinical symptoms, through mechanisms that differ among the two disorders. Several experimental studies have shown that noradrenaline modulates neuroinflammation, mainly by acting on microglia/astrocytes function. In the present review, after a brief introduction on the anatomy and physiology of LC, we provide an overview of experimental data supporting a pathogenetic role of LC degeneration in AD and PD. Then, we describe in detail experimental data, obtained in vitro and in vivo in animal models, which support a potential role of neuroinflammation in such a link, and the specific molecules (i.e., released cytokines, glial receptors, including pattern recognition receptors and others) whose expression is altered by LC degeneration and might play a key role in AD/PD pathogenesis. New imaging and biochemical tools have recently been developed in humans to estimate in vivo the integrity of LC, the degree of neuroinflammation, and pathology AD/PD biomarkers; it is auspicable that these will allow in the near future to test the existence of a link between LC-neuroinflammation and neurodegeneration directly in patients.

scholarly journals Targeting Mitochondrial Ion Channels to Fight Cancer

2018 ◽  
Vol 19 (7) ◽  
pp. 2060 ◽  
Author(s):  
Magdalena Bachmann ◽  
Roberto Costa ◽  
Roberta Peruzzo ◽  
Elena Prosdocimi ◽  
Vanessa Checchetto ◽  
...  
Keyword(s):  
Ion Channels ◽  
Tumor Development ◽  
Cancer Development ◽  
Cancer Growth ◽  
Altered Expression ◽  
New Frontier ◽  
Near Future

In recent years, several experimental evidences have underlined a new role of ion channels in cancer development and progression. In particular, mitochondrial ion channels are arising as new oncological targets, since it has been proved that most of them show an altered expression during tumor development and the pharmacological targeting of some of them have been demonstrated to be able to modulate cancer growth and progression, both in vitro as well as in vivo in pre-clinical mouse models. In this scenario, pharmacology of mitochondrial ion channels would be in the near future a new frontier for the treatment of tumors. In this review, we discuss the new advances in the field, by focusing our attention on the improvements in new drug developments to target mitochondrial ion channels.

scholarly journals “Cell-Free Synthetic Biology”: Synthetic Biology Meets Cell-Free Protein Synthesis

2019 ◽  
Vol 2 (4) ◽  
pp. 80
Author(s):  
Hong
Keyword(s):  
Protein Synthesis ◽  
Synthetic Biology ◽  
Biological Networks ◽  
Unnatural Amino Acids ◽  
Genetic Circuits ◽  
Free Protein ◽  
Protein Libraries ◽  
Cell Free Protein Synthesis

Since Nirenberg and Matthaei used cell-free protein synthesis (CFPS) to elucidate the genetic code in the early 1960s [1], the technology has been developed over the course of decades and applied to studying both fundamental and applied biology [2]. Cell-free synthetic biology integrating CFPS with synthetic biology has received attention as a powerful and rapid approach to characterize and engineer natural biological systems. The open nature of cell-free (or in vitro) biological platforms compared to in vivo systems brings an unprecedented level of control and freedom in design [3]. This versatile engineering toolkit has been used for debugging biological networks, constructing artificial cells, screening protein libraries, prototyping genetic circuits, developing biosensors, producing metabolites, and synthesizing complex proteins including antibodies, toxic proteins, membrane proteins, and novel proteins containing nonstandard (unnatural) amino acids. The Methods and Protocols “Cell-Free Synthetic Biology” Special Issue consists of a series of reviews, protocols, benchmarks, and research articles describing the current development and applications of cell-free synthetic biology in diverse areas. [...]

scholarly journals Incompatibility of DFHBI based fluorescent RNA aptamers with particular commercial cell-free expression systems

2021 ◽  
Author(s):  
Alexander J Speakman ◽  
Katherine E Dunn
Keyword(s):  
Expression System ◽  
In Vitro Transcription ◽  
Rna Aptamers ◽  
Free Expression ◽  
Reaction Products ◽  
Expression Systems ◽  
E Coli ◽  
Cell Free Expression System

Fluorescent RNA aptamers are an increasingly used tool for quantifying transcription and for visualising RNA interactions, both in vitro and in vivo. However when tested in the commercially available, E. coli extract based Expressway™ cell-free expression system, no fluorescence is detected. The same experimental setup is shown to successfully produce fluorescent RNA aptamers when tested in another buffer designed for in vitro transcription, and RNA purification of the Expressway™ reaction products show that transcription does occur, but does not result in a fluorescent product. In this paper we demonstrate the incompatibility of a narrow selection of RNA aptamers in one particular cell-free expression system, and consider that similar issues may arise with other cell-free expression systems, RNA aptamers, and their corresponding fluorophores.

scholarly journals Embryonated Eggs as an Alternative Infection Model To Investigate Aspergillus fumigatus Virulence

2010 ◽  
Vol 78 (7) ◽  
pp. 2995-3006 ◽  
Author(s):  
Ilse D. Jacobsen ◽  
Katharina Große ◽  
Silvia Slesiona ◽  
Bernhard Hube ◽  
Angela Berndt ◽  
...  
Keyword(s):  
Low Cost ◽  
Animal Experiments ◽  
Infection Model ◽  
Egg Survival ◽  
Chemokine Production ◽  
Rapid Construction ◽  
Infection Models ◽  
Invertebrate Models

ABSTRACT Infection models are essential tools for studying microbial pathogenesis. Murine models are considered the “gold standard” for studying in vivo infections caused by Aspergillus species, such as A. fumigatus. Recently developed molecular protocols allow rapid construction of high numbers of fungal deletion mutants, and alternative infection models based on cell culture or invertebrates are widely used for screening such mutants to reduce the number of rodents in animal experiments. To bridge the gap between invertebrate models and mice, we have developed an alternative, low-cost, and easy-to-use infection model for Aspergillus species based on embryonated eggs. The outcome of infections in the egg model is dose and age dependent and highly reproducible. We show that the age of the embryos affects the susceptibility to A. fumigatus and that increased resistance coincides with altered chemokine production after infection. The progress of disease in the model can be monitored by using egg survival and histology. Based on pathological analyses, we hypothesize that invasion of embryonic membranes and blood vessels leads to embryonic death. Defined deletion mutant strains previously shown to be fully virulent or partially or strongly attenuated in a mouse model of bronchopulmonary aspergillosis showed comparable degrees of attenuation in the egg model. Addition of nutrients restored the reduced virulence of a mutant lacking a biosynthetic gene, and variations of the infectious route can be used to further analyze the role of distinct genes in our model. Our results suggest that embryonated eggs can be a very useful alternative infection model to study A. fumigatus virulence and pathogenicity.

scholarly journals Effective Use of Linear DNA in Cell-Free Expression Systems

2021 ◽  
Vol 9 ◽  
Author(s):  
Megan A. McSweeney ◽  
Mark P. Styczynski
Keyword(s):  
Scientific Discovery ◽  
Free Expression ◽  
Genetic Circuit ◽  
Expression Systems ◽  
Linear Dna ◽  
Biological Phenomena ◽  
Wide Range ◽  
Research Tools

Cell-free expression systems (CFEs) are cutting-edge research tools used in the investigation of biological phenomena and the engineering of novel biotechnologies. While CFEs have many benefits over in vivo protein synthesis, one particularly significant advantage is that CFEs allow for gene expression from both plasmid DNA and linear expression templates (LETs). This is an important and impactful advantage because functional LETs can be efficiently synthesized in vitro in a few hours without transformation and cloning, thus expediting genetic circuit prototyping and allowing expression of toxic genes that would be difficult to clone through standard approaches. However, native nucleases present in the crude bacterial lysate (the basis for the most affordable form of CFEs) quickly degrade LETs and limit expression yield. Motivated by the significant benefits of using LETs in lieu of plasmid templates, numerous methods to enhance their stability in lysate-based CFEs have been developed. This review describes approaches to LET stabilization used in CFEs, summarizes the advancements that have come from using LETs with these methods, and identifies future applications and development goals that are likely to be impactful to the field. Collectively, continued improvement of LET-based expression and other linear DNA tools in CFEs will help drive scientific discovery and enable a wide range of applications, from diagnostics to synthetic biology research tools.

scholarly journals Role of the Furosemide Stress Test in Renal Injury Prognosis

2020 ◽  
Vol 21 (9) ◽  
pp. 3086 ◽  
Author(s):  
Armando Coca ◽  
Carmen Aller ◽  
Jimmy Reinaldo Sánchez ◽  
Ana Lucía Valencia ◽  
Elena Bustamante-Munguira ◽  
...  
Keyword(s):  
Kidney Disease ◽  
Clinical Decision Making ◽  
Low Cost ◽  
Stress Test ◽  
Clinical Decision ◽  
Clinical Routine ◽  
Patient Prognosis ◽  
Near Future ◽  
Published Research

Risk stratification and accurate patient prognosis are pending issues in the management of patients with kidney disease. The furosemide stress test (FST) has been proposed as a low-cost, fast, safe, and easy-to-perform test to assess tubular integrity, especially when compared to novel plasma and urinary biomarkers. However, the findings regarding its clinical use published so far provide insufficient evidence to recommend the generalized application of the test in daily clinical routine. Dosage, timing, and clinical outcomes of the FST proposed thus far have been significantly different, which further accentuates the need for standardization in the application of the test in order to facilitate the comparison of results between series. This review will summarize published research regarding the usefulness of the FST in different settings, providing the reader some insights about the possible implications of FST in clinical decision-making in patients with kidney disease and the challenges that research will have to address in the near future before widely applying the FST.

Mitochondrial matrix calcium ions regulate energy production in myocardium: A cytochemical study

1990 ◽  
Vol 48 (2) ◽  
pp. 166-167
Author(s):  
W.A. Jacob ◽  
R. Hertsens ◽  
A. Van Bogaert ◽  
M. De Smet
Keyword(s):  
Energy Metabolism ◽  
Calcium Ions ◽  
Energy Production ◽  
Regulation Of Respiration ◽  
Free Calcium ◽  
Mitochondrial Matrix ◽  
Cytochemical Study ◽  
Nmr Techniques

In the past most studies of the control of energy metabolism focus on the role of the phosphorylation potential ATP/ADP.Pi on the regulation of respiration. Studies using NMR techniques have demonstrated that the concentrations of these compounds for oxidation phosphorylation do not change appreciably throughout the cardiac cycle and during increases in cardiac work. Hence regulation of energy production by calcium ions, present in the mitochondrial matrix, has been the object of a number of recent studies.Three exclusively intramitochondnal dehydrogenases are key enzymes for the regulation of oxidative metabolism. They are activated by calcium ions in the low micromolar range. Since, however, earlier estimates of the intramitochondnal calcium, based on equilibrium thermodynamic considerations, were in the millimolar range, a physiological correlation was not evident. The introduction of calcium-sensitive probes fura-2 and indo-1 made monitoring of free calcium during changing energy metabolism possible. These studies were performed on isolated mitochondria and extrapolation to the in vivo situation is more or less speculative.

A technique for protecting delicate specimens during processing

1989 ◽  
Vol 47 ◽  
pp. 982-983
Author(s):  
R.J. Mount ◽  
R.V. Harrison
Keyword(s):  
Basilar Membrane ◽  
Low Cost ◽  
Organ Of Corti ◽  
Region Of Interest ◽  
Sensory Epithelium ◽  
Physical Damage ◽  
Air Drying ◽  
Specimen Handling ◽  
Two Component

The sensory end organ of the ear, the organ of Corti, rests on a thin basilar membrane which lies between the bone of the central modiolus and the bony wall of the cochlea. In vivo, the organ of Corti is protected by the bony wall which totally surrounds it. In order to examine the sensory epithelium by scanning electron microscopy it is necessary to dissect away the protective bone and expose the region of interest (Fig. 1). This leaves the fragile organ of Corti susceptible to physical damage during subsequent handling. In our laboratory cochlear specimens, after dissection, are routinely prepared by the O-T- O-T-O technique, critical point dried and then lightly sputter coated with gold. This processing involves considerable specimen handling including several hours on a rotator during which the organ of Corti is at risk of being physically damaged. The following procedure uses low cost, readily available materials to hold the specimen during processing ,preventing physical damage while allowing an unhindered exchange of fluids.Following fixation, the cochlea is dehydrated to 70% ethanol then dissected under ethanol to prevent air drying. The holder is prepared by punching a hole in the flexible snap cap of a Wheaton vial with a paper hole punch. A small amount of two component epoxy putty is well mixed then pushed through the hole in the cap. The putty on the inner cap is formed into a “cup” to hold the specimen (Fig. 2), the putty on the outside is smoothed into a “button” to give good attachment even when the cap is flexed during handling (Fig. 3). The cap is submerged in the 70% ethanol, the bone at the base of the cochlea is seated into the cup and the sides of the cup squeezed with forceps to grip it (Fig.4). Several types of epoxy putty have been tried, most are either soluble in ethanol to some degree or do not set in ethanol. The only putty we find successful is “DUROtm MASTERMENDtm Epoxy Extra Strength Ribbon” (Loctite Corp., Cleveland, Ohio), this is a blue and yellow ribbon which is kneaded to form a green putty, it is available at many hardware stores.

Sign in / Sign up

Export Citation Format

Share Document