Cell-Free Synthetic Biology Platform for Engineering Synthetic Biological Circuits and Systems

Synthetic biology brings engineering disciplines to create novel biological systems for biomedical and technological applications. The substantial growth of the synthetic biology field in the past decade is poised to transform biotechnology and medicine. To streamline design processes and facilitate debugging of complex synthetic circuits, cell-free synthetic biology approaches has reached broad research communities both in academia and industry. By recapitulating gene expression systems in vitro, cell-free expression systems offer flexibility to explore beyond the confines of living cells and allow networking of synthetic and natural systems. Here, we review the capabilities of the current cell-free platforms, focusing on nucleic acid-based molecular programs and circuit construction. We survey the recent developments including cell-free transcription– translation platforms, DNA nanostructures and circuits, and novel classes of riboregulators. The links to mathematical models and the prospects of cell-free synthetic biology platforms will also be discussed.11Yscopu

Investigation on Health Effects of an Abandoned Metal Mine

To investigate potential health risks associated with exposure to metals from an abandoned metal mine, the authors studied people living near an abandoned mine (n=102) and control groups (n=149). Levels of cadmium, copper, arsenic, lead, and zinc were measured in the air, soil, drinking water, and agricultural products. To assess individual exposure, biomarkers of each metal in blood and urine were measured. β2-microglobulin, α1-microglobulin, and N-acetyl-beta-glucosaminidase and bone mineral density were measured. Surface soil in the study area showed 2-10 times higher levels of metals compared to that of the control area. Metal concentrations in the groundwater and air did not show any notable differences between groups. Mean concentrations of cadmium and copper in rice and barley from the study area were significantly higher than those of the control area (p<0.05). Geometric means of blood and urine cadmium in the study area were 2.9 µg/L and 1.5 µg/g Cr, respectively, significantly higher than those in the control area (p<0.05). There were no differences in the levels of urinary markers of early kidney dysfunction and bone mineral density. The authors conclude that the residents near the abandoned mine were exposed to higher levels of metals through various routes

Cellular Computational Logic Using Toehold Switches

The development of computational logic that carries programmable and predictable features is one of the key requirements for next-generation synthetic biological devices. Despite considerable progress, the construction of synthetic biological arithmetic logic units presents numerous challenges. In this paper, utilizing the unique advantages of RNA molecules in building complex logic circuits in the cellular environment, we demonstrate the RNA-only bitwise logical operation of XOR gates and basic arithmetic operations, including a half adder, a half subtractor, and a Feynman gate, in Escherichia coli. Specifically, de-novo-designed riboregulators, known as toehold switches, were concatenated to enhance the functionality of an OR gate, and a previously utilized antisense RNA strategy was further optimized to construct orthogonal NIMPLY gates. These optimized synthetic logic gates were able to be seamlessly integrated to achieve final arithmetic operations on small molecule inputs in cells. Toehold-switch-based ribocomputing devices may provide a fundamental basis for synthetic RNA-based arithmetic logic units or higher-order systems in cells11Ysciescopu

Cellular Arithmetic Logic Unit Using Toehold Switches

Since the demonstration of primitive studies on molecular computing over two decades ago, modern synthetic biologists are now seeking for more complex artificial cellular functions. In this regard, RNA is one of promising materials for synthetic circuit due to its fully designable Watson-Crick base pairing and various structure prediction bioinformatic tools. While several arithmetic logics are implemented using diverse components [1,2], RNA-only operation has not yet been achieved. Here, we show RNA-only binary operation of XOR gate and cellular arithmetic logic operation, specifically half-adder. By the integration of orthogonal NIMPLY gates developed from de-novo-designed toehold switches [3], we can successfully demonstrate XOR gate which have higher dynamic range compared to conventional systems. To achieve comprehensive screening of functional antisense RNAs, we employed automatic design pipeline for selected toehold switch pairs. Then, basic binary calculator was constructed based on our XOR gate by combining with AND gate. We have further planned to build more sophisticated molecular computing devices, such as Neural network, by using multi-input multi-output features of binary calculators.2

Detection of pks Island mRNAs Using Toehold Sensors in Escherichia coli

Synthetic biologists have applied biomolecular engineering approaches toward the goal of novel biological devices and have shown progress in diverse areas of medicine and biotechnology. Especially promising is the application of synthetic biological devices towards a novel class of molecular diagnostics. As an example, a de-novo-designed riboregulator called toehold switch, with its programmability and compatibility with field-deployable devices showed promising in vitro applications for viral RNA detection such as Zika and Corona viruses. However, the in vivo application of high-performance RNA sensors remains challenging due to the secondary structure of long mRNA species. Here, we introduced &lsquo;Helper RNAs&rsquo; that can enhance the functionality of toehold switch sensors by mitigating the effect of secondary structures around a target site. By employing the helper RNAs, previously reported mCherry mRNA sensor showed improved fold-changes in vivo. To further generalize the Helper RNA approaches, we employed automatic design pipeline for toehold sensors that target the essential genes within the pks island, an important target of biomedical research in connection with colorectal cancer. The toehold switch sensors showed fold-changes upon the expression of full-length mRNAs that apparently depended sensitively on the identity of the gene as well as the predicted local structure within the target region of the mRNA. Still, the helper RNAs could improve the performance of toehold switch sensors in many instances, with up to 10-fold improvement over no helper cases. These results suggest that the helper RNA approaches can further assist the design of functional RNA devices in vivo with the aid of the streamlined automatic design software developed here. Further, our solutions for screening and stabilizing single-stranded region of mRNA may find use in other in vivo mRNA-sensing applications such as cas13 crRNA design, transcriptome engineering, and trans-cleaving ribozymes

Detection of pks Island mRNAs Using Toehold Sensors in Escherichia coli

Synthetic biologists have applied biomolecular engineering approaches toward the goal of novel biological devices and have shown progress in diverse areas of medicine and biotechnology. Especially promising is the application of synthetic biological devices towards a novel class of molecular diagnostics. As an example, a de-novo-designed riboregulator called toehold switch, with its programmability and compatibility with field-deployable devices showed promising in vitro applications for viral RNA detection such as Zika and Corona viruses. However, the in vivo application of high-performance RNA sensors remains challenging due to the secondary structure of long mRNA species. Here, we introduced ‘Helper RNAs’ that can enhance the functionality of toehold switch sensors by mitigating the effect of secondary structures around a target site. By employing the helper RNAs, previously reported mCherry mRNA sensor showed improved fold-changes in vivo. To further generalize the Helper RNA approaches, we employed automatic design pipeline for toehold sensors that target the essential genes within the pks island, an important target of biomedical research in connection with colorectal cancer. The toehold switch sensors showed fold-changes upon the expression of full-length mRNAs that apparently depended sensitively on the identity of the gene as well as the predicted local structure within the target region of the mRNA. Still, the helper RNAs could improve the performance of toehold switch sensors in many instances, with up to 10-fold improvement over no helper cases. These results suggest that the helper RNA approaches can further assist the design of functional RNA devices in vivo with the aid of the streamlined automatic design software developed here. Further, our solutions for screening and stabilizing single-stranded region of mRNA may find use in other in vivo mRNA-sensing applications such as cas13 crRNA design, transcriptome engineering, and trans-cleaving ribozymes.11Ysciescopu

Early snapshot on exposure to environmental chemicals among Korean adults-results of the first Korean National Environmental Health Survey (2009-2011)

Background: Chemical exposure may cause serious adverse health effects. Under the Environmental Health Act in Korea, the first national survey, Korean National Environmental Health Survey (KoNHES), was conducted, and dozens of major environmental chemicals were measured among Korean adults between 2009 and 2011. Objectives: To understand the levels of exposure to environmental chemicals among general Korean adults population, and to identify factors that may influence the levels of exposure. Methods: The blood and urine samples were collected from Korean adults with 19 years of age and older (n = 6311), and were analyzed for a number of environmental chemicals including metals, bisphenol A (BPA), and metabolites of polycyclic aromatic hydrocarbons (PAHs), phthalates, a pyrethroid insecticide (3-PBA) and several volatile organic carbons (VOCs). Demographic and other behavioral factors were asked through questionnaire. Results: Among general Korean adults, mercury, arsenic, cadmium, and metabolite of DBP (MnBP) were higher than those reported in other nationwide surveys of Canada, Germany, and USA. Most target chemicals except for MEOHP and 3-PBA showed significant differences on the exposure levels by sex Conclusion: This is the first nationwide reconnaissance on exposure to environmental chemicals among general Korean population. Chemicals with high occurrence level will be further investigated to identify exposure sources and possible health outcomes, and eventually to develop measures to mitigate exposure. (C) 2016 Published by Elsevier GmbH.OAIID:RECH_ACHV_DSTSH_NO:T201635601RECH_ACHV_FG:RR00200001ADJUST_YN:EMP_ID:A002120CITE_RATE:4.643DEPT_NM:환경보건학과EMAIL:[email protected]_YN:YN

Dosimetric evaluation of respiratory gated volumetric modulated arc therapy for lung stereotactic body radiation therapy using 3D printing technology.

PurposeThis study aimed to evaluate the dosimetric accuracy of respiratory gated volumetric modulated arc therapy (VMAT) for lung stereotactic body radiation therapy (SBRT) under simulation conditions similar to the actual clinical situation using patient-specific lung phantoms and realistic target movements.MethodsSix heterogeneous lung phantoms were fabricated using a 3D-printer (3DISON, ROKIT, Seoul, Korea) to be dosimetrically equivalent to actual target regions of lung SBRT cases treated via gated VMAT. They were designed to move realistically via a motion device (QUASAR, Modus Medical Devices, Canada). Using the lung phantoms and a homogeneous phantom (model 500-3315, Modus Medical Devices), film dosimetry was performed with and without respiratory gating for VMAT delivery (TrueBeam STx; Varian Medical Systems, Palo Alto, CA, USA). The measured results were analyzed with the gamma passing rates (GPRs) of 2%/1 mm criteria, by comparing with the calculated dose via the AXB and AAA algorithms of the Eclipse Treatment Planning System (version 10.0.28; Varian Medical Systems).ResultsGPRs were greater than the acceptance criteria 80% for all film measurements with the stationary and homogeneous phantoms in conventional QAs. Regardless of the heterogeneity of phantoms, there were no significant differences (p > 0.05) in GPRs obtained with and without target motions; the statistical significance (p = 0.031) was presented between both algorithms under the utilization of heterogeneous phantoms.ConclusionsDosimetric verification with heterogeneous patient-specific lung phantoms could be successfully implemented as the evaluation method for gated VMAT delivery. In addition, it could be dosimetrically confirmed that the AXB algorithm improved the dose calculation accuracy under patient-specific simulations using 3D printed lung phantoms