A grand challenge in synthetic biology is to use our current knowledge of RNA science perform the automatic engineering completely sequences encoding functional RNAs living cells. We report here a fully automated design methodology and experimental validation interaction circuits working cellular environment. The computational algorithm, based on physicochemical model, produces novel by exploring space possible compatible with predefined structures. tested Escherichia coli designing several...

Viral infections in plants threaten food security. Thus, simple and effective methods for virus detection are required to adopt early measures that can prevent spread. However, current based on the amplification of viral genome by polymerase chain reaction (PCR) require laboratory conditions. Here, we exploited CRISPR-Cas12a CRISPR-Cas13a/d systems detect three RNA viruses, namely, Tobacco mosaic virus, etch Potato X, Nicotiana benthamiana plants. We applied system DNA amplicons generated...

Abstract Motivation: The biological solution for synthesis or remediation of organic compounds using living organisms, particularly bacteria and yeast, has been promoted because the cost reduction with respect to non-living chemical approach. In that way, computational frameworks can profit from previous knowledge stored in large databases compounds, enzymes reactions. addition, cell behavior be studied by modeling cellular context. Results: We have implemented a Monte Carlo algorithm...

The genomes of plant viruses have limited coding capacity, and to complete their infectious cycles, viral factors must target, direct or indirectly, many host elements. However, the interaction networks between are poorly understood. genus Potyvirus is largest group plus-strand RNA infecting plants. Potyviral nuclear inclusion a (NIa) plays roles during infection. NIa polyprotein consisting two domains, protein genome-linked (VPg) protease (NIaPro), separated by an inefficiently utilized...

Volatile organic compounds (VOCs) are major determinants of fruit flavor, a primary objective in tomato breeding. A recombinant inbred line (RIL) population consisting 169 lines derived from cross between Solanum lycopersicum and red-fruited wild species pimpinellifolium accession (SP) was characterized for VOCs three different seasons. Correlation hierarchical cluster analyses were performed on the 52 identified, providing tool putative assignation individual to metabolic pathways....

Abstract Viruses are obligate intracellular parasites that have co-evolved with their hosts to establish an intricate network of protein–protein interactions. Here, we followed a high-throughput yeast two-hybrid screening identify 378 novel interactions between turnip mosaic virus (TuMV) and its natural host Arabidopsis thaliana . We identified the RNA-dependent RNA polymerase NIb as viral protein largest number contacts, including key salicylic acid-dependent transcription regulators....

Understanding the molecular mechanisms plants have evolved to adapt their biological activities a constantly changing environment is an intriguing question and one that requires systems biology approach. Here we present network analysis of genome-wide expression data combined with reverse-engineering modeling dissect transcriptional control Arabidopsis thaliana. The regulatory inferred by using assembly microarray containing steady-state RNA levels from several growth conditions,...

Synthetic biology uses modeling to facilitate the design of new genetic constructions. In particular, it is utmost importance model reaction cellular chassis when expressing heterologous systems. We constructed a mathematical for response bacterial cell under expression. For this, we relied on previous characterization growth-rate dependence resource availability (in this case, DNA and RNA polymerases ribosomes). Accordingly, estimated maximum capacities expression be 46% total 33% protein....

A complex network of cellular receptors, RNA targeting pathways, and small-molecule signaling provides robust plant immunity tolerance to viruses. To maximize their fitness, viruses must evolve control mechanisms balance host immune evasion plant-damaging effects. The genus Potyvirus comprises characterized by genomes that encode large polyproteins led the P1 protease. autoinhibitory domain controls polyprotein processing, release a downstream functional RNA-silencing suppressor, viral...

MiRNAs have emerged as key regulators of stress response in plants, suggesting their potential candidates for knock-in/out to improve tolerance agricultural crops. Although diverse assays been performed, systematic and detailed studies miRNA expression function during exposure multiple environments crops are limited. Here, we present such pioneering analysis melon plants seven biotic abiotic conditions. Deep-sequencing computational approaches identified twenty-four known miRNAs whose was...

Recurrent disease outbreaks caused by different viruses, including the novel respiratory virus SARS-CoV-2, are challenging our society at a global scale; so versatile detection methods would enable calculated and faster response. Here, we present nucleic acid strategy based on CRISPR-Cas9, whose mode of action relies strand displacement rather than collateral catalysis, using Streptococcus pyogenes Cas9 nuclease. Given preamplification process, suitable molecular beacon interacts with...

Development of a novel framework to regulate translation by repurposing the CRISPR-Cas13 immune system, exploiting cell-free expression system for prototyping.

In public health emergencies or in resource-constrained settings, laboratory-based diagnostic methods, such as RT-qPCR, need to be complemented with accurate, rapid, and accessible approaches increase testing capacity, this will translate into better outcomes disease prevention management. Here, we develop an original nucleic acid detection platform by leveraging the CRISPR-Cas9 lateral flow immunochromatography technologies. combination isothermal amplification that runs a biotinylated...

Copper (Cu) deficiency compromises plant growth and limits crop production. Plants respond to Cu scarcity by activating the expression of several microRNAs, known as Cu-miRNAs, which degrade mRNAs from various cuproproteins conserve Cu. like most miRNAs, associate with ARGONAUTE 1 (AGO1), primary effector protein miRNA-mediated gene silencing pathways, whose function is typically modulated interacting proteins acting cofactors. However, how AGO1 regulated functions under remains unknown....

Reporter systems are widely used to study biomolecular interactions and processes in vivo, representing one of the basic tools characterize synthetic regulatory circuits. Here, we developed a method that enables monitoring RNA–protein through reporter system bacteria with high temporal resolution. For this, Real-Time Protein Expression Assay (RT-PEA) technology for real-time fluorescent protein, while having growing on solid media. Experimental results were analyzed by fitting three-variable...

Abstract Motivation: The rational design of biological networks with prescribed functions is limited to gene circuits a few genes. Larger involve complex interactions many parameters and the use automated computational tools can be very valuable. We propose new tool transcriptional targeted behavior that could used better understand principles genetic circuits. Results: have implemented Simulated Annealing optimization algorithm explores throughout space transcription obtain specific...

Tobacco etch potyvirus (TEV) has been extensively used as model system for the study of positive-sense RNA virus infecting plants. TEV ability to infect Arabidopsis thaliana varies among ecotypes. In this study, changes in gene expression A. ecotype Ler infected with have explored using long-oligonucleotide arrays. is a susceptible host that allows systemic movement, although viral load low and syndrome induced ranges from asymptomatic mild. Gene profiles were monitored whole plants 21 days...

Synthetic regulatory networks with prescribed functions are engineered by assembling a reduced set of functional elements. We could also assemble them computationally if the mathematical models those elements were predictive enough in different genetic contexts. Only after achieving this will we have libraries biological parts able to provide dynamical behaviors for most circuits constructed them. thus need tools that can automatically explore contexts, addition being use such design novel...

The engineering of synthetic gene networks has mostly relied on the assembly few characterized regulatory elements using rational design principles. It is outmost importance to analyze scalability and limits such a workflow. To capabilities libraries elements, we have developed first automated approach that combines search genotype space associated given phenotypic behavior. Herein, calculated designability dynamical functions obtained from circuits assembled with genetic library. By...

Small RNAs (sRNAs) can operate as regulatory agents to control protein expression by interaction with the 5′ untranslated region of mRNA. We have developed a physicochemical framework, relying on base pair energies, design multi-state sRNA devices solving an optimization problem objective function accounting for stability transition and final intermolecular states. Contrary analysis reaction kinetics ensemble sRNAs, we solve inverse finding sequences satisfying targeted reactions. show here...