In vitro molecular pattern classification via DNA-based weighted-sum operation

Recent progress in molecular computation suggests the possibility of pattern classification in vitro. Weighted sum is a primitive operation required by many pattern classification problems. Here we present a DNA-based molecular computation method for implementing the weighted-sum operation and its use for molecular pattern classification in a test tube. The weights of the classifier are encoded as the mixing ratios of the differentially labeled probe DNA molecules, which are competitively hybridized with the input-encoding target molecules to compute the decision boundary of classification. The computation result is detected by fluorescence signals. We experimentally verify the underlying weight encoding scheme and demonstrate successful discrimination of two-group labels of synthetic DNA mixture patterns. The method can be used for direct computation on biomolecular data in a liquid state.     

Phase separation in amino acid mixtures is governed by composition

Macromolecular phase separation has recently come to immense prominence as it is central to the formation of membraneless organelles, leading to a new paradigm of cellular organization. This type of phase transition, often termed liquid-liquid phase separation (LLPS), is mediated by molecular interactions between biomolecules, including nucleic acids and both ordered and disordered proteins. In the latter case, the separation between protein-dense and -dilute phases is often interpreted using models adapted from polymer theory. Specifically, the “stickers and spacers” model proposes that the formation of condensate-spanning networks in protein solutions originates from the interplay between two classes of residues and that the main determinants for phase separation are multivalency and sequence patterning. The duality of roles of stickers (aromatics like Phe and Tyr) and spacers (Gly and polar residues) may apply more broadly in protein-like mixtures, and the presence of these two types of components alone may suffice for LLPS to take place. In order to explore this hypothesis, we use atomistic molecular dynamics simulations of capped amino acid residues as a minimal model system. We study the behavior of pure amino acids in water for three types of residues corresponding to the spacer and sticker categories and of their multicomponent mixtures. In agreement with previous observations, we find that the spacer-type amino acids fail to phase separate on their own, while the sticker is prone to aggregation. However, ternary amino acid mixtures involving both types of amino acids phase separate into two phases that retain intermediate degrees of compaction and greater fluidity than sticker-only condensates. Our results suggest that LLPS is an emergent property of amino acid mixtures determined by composition.     

Approaching mathematical model of the immune network based DNA Strand Displacement system

One biggest obstacle in molecular programming is that there is still no direct method to compile any existed mathematical model into biochemical reaction in order to solve a computational problem. In this paper, the implementation of DNA Strand Displacement system based on nature-inspired computation is observed. By using the Immune Network Theory and Chemical Reaction Network, the compilation of DNA-based operation is defined and the formulation of its mathematical model is derived. Furthermore, the implementation on this system is compared with the conventional implementation by using silicon-based programming. From the obtained results, we can see a positive correlation between both. One possible application from this DNA-based model is for a decision making scheme of intelligent computer or molecular robot.     

The cell as the smallest DNA-based molecular computer

The pioneering work of Adleman (1994) demonstrated that DNA molecules in test tubes can be manipulated to perform a certain type of mathematical computation. This has stimulated a theoretical interest in the possibility of constructing DNA-based molecular computers. To gauge the practicality of realizing such microscopic computers, it was thought necessary to learn as much as possible from the biology of the living cell--presently the only known DNA-based molecular computer in existence. Here the recently developed theoretical model of the living cell (the Bhopalator) and its associated theories (e.g. cell language), principles, laws and concepts (e.g. conformons, IDS's) are briefly reviewed and summarized in the form of a set of five laws of 'molecular semiotics' (synonyms include 'microsemiotics', 'cellular semiotics', or 'cytosemiotics') the study of signs mediating measurement, computation, and communication on the cellular and molecular levels. Hopefully, these laws will find practical applications in designing DNA-based computing systems.     

Fish as predators and prey: DNA-based assessment of their role in food webs

Fish are both consumers and prey, and as such part of a dynamic trophic network. Measuring how they are trophically linked, both directly and indirectly, to other species is vital to comprehend the mechanisms driving alterations in fish communities in space and time. Moreover, this knowledge also helps to understand how fish communities respond to environmental change and delivers important information for implementing management of fish stocks. DNA-based methods have significantly widened our ability to assess trophic interactions in both marine and freshwater systems and they possess a range of advantages over other approaches in diet analysis. In this review we provide an overview of different DNA-based methods that have been used to assess trophic interactions of fish as consumers and prey. We consider the practicalities and limitations, and emphasize critical aspects when analysing molecular derived trophic data. We exemplify how molecular techniques have been employed to unravel food web interactions involving fish as consumers and prey. In addition to the exciting opportunities DNA-based approaches offer, we identify current challenges and future prospects for assessing fish food webs where DNA-based approaches will play an important role.     

Molecular approaches to understanding mycorrhizal symbioses

Molecular analyses of plant–microbe interactions have become common place in the last two decades. Although there are philosophical considerations about the application of a reductionist approach to some areas of research, the collaborative interface (e.g. molecular ecology) can provide specialised insight to the generalist, whilst adding broader relevance to the research of the specialist. However, the expense of this discipline has tended to restrict research to work on model host–microbe interactions. Molecular techniques were embraced early on by a few pioneers from the field of mycorrhizal research. Despite some high profile research, the number of molecular mycorrhizal publications has only recently begun to escalate. However the extent of literature now has exceeded the capacity for a comprehensive short review. In this paper we will briefly discuss the use of model species for molecular research and explore the range of questions that are being addressed using molecular techniques, whilst minimising use of specific jargon, to maximise the usefulness of this review to a non specialist audience. Our primary focus is on arbuscular mycorrhizal symbiosis, to complement the papers by Tagu et al., Podila et al. and Chalot et al. (all this volume), who have addressed aspects of research on ectomycorrhizal symbioses. Here we include specific citations from research groups around the world, along with reference to more detailed reviews, to provide a taste of the current excitement in this fundamental and rapidly evolving research field.     

The anchorosome, a special chromatin granule for the anchorage of the interphase chromosome to the nuclear envelope

Peripheral chromatin granules bound to the nuclear envelope of rat liver nuclei have been further investigated. Judging by the results of Staphylococcal nuclease digestion of nuclei and electron microscopical observations, the peripheral granules have nucleosomal organization. As shown by ultraviolet radiation DNA-protein cross-linkage, the histone-like proteins present in the peripheral chromatin instead of histone H1 (Fais et al., 1982) are in close contact with DNA. The peripheral chromatin contains a DNA firmly bound to the lamina. This DNA, resistant to extraction in high salt, heparin and SDS, is protected against a DNase attack since, as shown by DNA electrophoresis data, high molecular weight molecules (up to 20 kbas) are still present in the lamina residue. However, the high molecular weight DNA disappeared if the nuclear envelope fraction was again DNase-digested after high salt treatment. Altogether, the data of the previous (Fais et al., 1982; Prusov et al., 1980: Prusov et al., 1982) and the present investigations demonstrate that the peripheral chromatin granules are endowed with properties which distinguish them from the bulk chromatin and account for the chromosome bond to the nuclear envelope during interphase. This is why we suggest the term "anchorosome" for the peripheral protein granule attached to the nuclear envelope.     

Quantitative prediction of 3D solution shape and flexibility of nucleic acid nanostructures

DNA nanotechnology enables the programmed synthesis of intricate nanometer-scale structures for diverse applications in materials and biological science. Precise control over the 3D solution shape and mechanical flexibility of target designs is important to achieve desired functionality. Because experimental validation of designed nanostructures is time-consuming and cost-intensive, predictive physical models of nanostructure shape and flexibility have the capacity to enhance dramatically the design process. Here, we significantly extend and experimentally validate a computational modeling framework for DNA origami previously presented as CanDo [Castro,C.E., Kilchherr,F., Kim,D.-N., Shiao,E.L., Wauer,T., Wortmann,P., Bathe,M., Dietz,H. (2011) A primer to scaffolded DNA origami. Nat. Meth., 8, 221-229.]. 3D solution shape and flexibility are predicted from basepair connectivity maps now accounting for nicks in the DNA double helix, entropic elasticity of single-stranded DNA, and distant crossovers required to model wireframe structures, in addition to previous modeling (Castro,C.E., et al.) that accounted only for the canonical twist, bend and stretch stiffness of double-helical DNA domains. Systematic experimental validation of nanostructure flexibility mediated by internal crossover density probed using a 32-helix DNA bundle demonstrates for the first time that our model not only predicts the 3D solution shape of complex DNA nanostructures but also their mechanical flexibility. Thus, our model represents an important advance in the quantitative understanding of DNA-based nanostructure shape and flexibility, and we anticipate that this model will increase significantly the number and variety of synthetic nanostructures designed using nucleic acids.     

105 Activity of DNA ligase on substrates containing non-canonical structures

The expansion of trinucleotide repeat tracts (e.g. (CAG)_n tracts) has been shown to contribute to genomic instability and has been implicated in the pathogenesis of several neurodegenerative diseases, including Huntington’s Disease and Fragile X syndrome (Kovtun et al., 2008). While the molecular mechanism of this expansion is unknown, the ability of trinucleotide repeat sequences to form non-canonical secondary structures, such as hairpins, has been implicated as a multifaceted source of error (Gacy et al., 1995). Non-canonical DNA secondary structures have been shown to impact the action of enzymes in the base excision repair (BER) pathway, by which oxidatively damaged bases are removed. More specifically, there is evidence that trinucleotide repeat-containing DNA mistakenly enters long-patch BER, which can potentially lead to the incorporation of extra nucleobases by DNA polymerase (Jarem et al., 2011). The final enzyme in the BER pathway is DNA Ligase, which catalyses the formation of a phosphodiester bond to seal a nick site (Taylor et al., 2011). When extra nucleotides have been added during an erroneous long-patch BER process, the action of DNA ligase may expand the repeat tract by incorporating these additional bases into duplex DNA. In this study, DNA constructs containing (CAG)_n hairpins at various distances from a nick site are used to investigate the ability of DNA Ligase to ligate substrates containing non-canonical secondary structure back into duplex DNA.     

The bounded complexity of DNA computing

This paper proposes a new approach to analyzing DNA-based algorithms in molecular computation. Such protocols are characterized abstractly by: encoding, tube operations and extraction. Implementation of these approaches involves encoding in a multiset of molecules that are assembled in a tube having a number of physical attributes. The physico-chemical state of a tube can be changed by a prescribed number of elementary operations. Based on realistic definitions of these elementary operations, we define complexity of a DNA-based algorithm using the physico-chemical property of each operation. We show that new algorithms for Hamiltonian path are about twice as efficient as Adleman's original one and that a recent algorithm for Max-Clique provides a similar increase in efficiency. Consequences of this approach to tube complexity and DNA computing are discussed.     

Confounding social and mating systems predictably lead to biased results when examining the evolution of cooperative breeding in cichlids: A response to Tanaka et al.

In 2017, we demonstrated that transitions to cooperative breeding in Lamprologine cichlid fishes were not related to a species’ social mating system (Dey et al. 2017. Nature Ecology & Evolution, 1 , 137). This contrasted previous evidence that monogamy (and a low degree of promiscuity) promoted transitions to cooperative breeding in other taxa. Recently, Tanaka et al. (2018. Ethology, 124 , 777–789) critiqued our study and argued that a re‐analysis of the data shows transitions to cooperative breeding are promoted by non‐monogamous mating systems. Here, we show that Tanaka et al.'s critique contains numerous inaccuracies. In addition, we show that the results put forth by Tanaka et al. emerge only under the extreme scenario in which all cooperative breeding species are classified as non‐monogamous, which we argue arises because Tanaka et al. confound social systems and mating systems. While we agree that there is uncertainty regarding the mating system of some Lamprologine species, we argue this uncertainty was sufficiently addressed through the extensive sensitivity analyses conducted in our original study.     

DNA engineering and its application to nanotechnology.

The combination of branched DNA molecules and 'sticky' ends creates a powerful molecular assembly kit for structural DNA nanotechnology. Polyhedra, complex topological objects, a nanomechanical device and two-dimensional arrays with programmable surface features have already been produced in this way. Future applications range from macromolecular crystallography and new materials to molecular electronics and DNA-based computation.     

The correlation between development of atypical bisexual flowers and phylogeny in the Aroideae (Araceae)

 In the intermediate zone of the inflorescence of genera of Aroideae one can find flowers with male and female characteristics. Until now, two types of developmental sequences of atypical bisexual flowers (ABFs) have been recognized: the Philodendron type and the Cercestis type. In the Philodendron type, bisexual flowers generally consist of functional carpels and staminodes inserted on the same whorl. In the Cercestis type, the gynoecium and stamens are inserted on two different whorls. These different ontogenetic patterns represent two different pathways in the evolution of unisexual flowers in this subfamily. A molecular phylogenetic analysis of 33 genera of Araceae, based on the chloroplast trnL intron and trnL–F intergenic spacer sequences was carried out. We use this phylogenetic analysis and those published by French et al. (1995) and Mayo et al. (1997) to examine the distribution of the two types of ABFs in selected genera. Our results suggest that the two developmental patterns of ABFs in Aroideae sensu Mayo et al. (1997) do not correspond to two separate evolutionary lineages but rather are more or less consistent within clades. Although this new molecular phylogeny does not include all aroid genera, it corroborates in general, at the subfamily level, the molecular analysis of French et al. (1995) based on chloroplast DNA restriction site data and the analysis of Mayo et al. (1997) based on morphological and anatomical data. Received March 15, 2001 Accepted October 11, 2001     

Optimizing conditions for DNA isolation fromPinus radiata

Summary Genomic DNA was isolated fromin vitro Pinus radiata seedling with five DNA isolation protocols commonly used for pines. The methods described by Jobes et al. (1995) and Nelson et al. (1994) utilize sodium dodecyl sulfate, whereas those of Murray and Thompson (1980), Doyle and Doyle (1990), and Devey et al. (1996) use cetyltrimethyl ammonium bromide for cell lysis. The quality and quantity of the isolated DNA was measured and compared. Lithium chloride was found to be more effective than RNase for minimizing the amount of RNA present in the solution. Protocols described by Jobes et al. (1995) and Devey et al. (1996) yielded a large quantity of pure DNA which was suitable for restriction enzyme digestion and polymerase chain reaction amplification. With these methods, 37 to 79 μg of DNA with an A_260/280 ratio between 1.7 and 1.9 was obtained from 1 g ofPinus radiata seedlings grownin vitro.     

44 Circular dichroism (CD) studies of the interaction between G-quadrulexes/I-motif with saffron secondary active metabolites

Telomeric DNA contains some unique secondary structures, such as G-quadruplex and I-motif. These structures may be stabilized or changed by binding to specific proteins or small molecules. In continuation of our previous studies on the interaction between crocin and crocetin, as the natural C20 carotenoids, and picrocrocin and safranal, as the natural monoterpene aldehydes, obtained from saffron and DNA (Bathaie et al., 2007; Hoshyar et al., 2008), herein, we report the in vitro effect of these saffron metabolites on the mentioned structures (Hoshyar et al., 2012). Circular dichroism (CD) data indicate that crocetin has higher affinity for these structures. Safranal and crocin induce little change in the I-motif and G-quadruplex, respectively. The molecular docking confirms the experimental data and indicates the minor groove binding of ligands with G-quadruplex. Effects of these ligands on the stability of these structures is studied using some other techniques and determination of thermodynamic parameters.     

A review of heavy metal cation binding to deoxyribonucleic acids for the creation of chemical sensors

Various human activities lead to the pollution of ground, drinking, and wastewater with toxic metals. It is well known that metal ions preferentially bind to DNA phosphate backbones or DNA nucleobases, or both. Foreman et al. (Environ Toxicol Chem 30(8):1810–1818, 2011) reported the use of a DNA-dye based assay suitable for use as a toxicity test for potable environmental water. They compared the results of this test with the responses of live-organism bioassays. The DNA-based demonstrated that the loss of SYBR Green I fluorescence dye bound to calf thymus DNA was proportional to the toxicity of the water sample. However, this report raised questions about the mechanism that formed the basis of this quasi-quantitatively test. In this review, we identify the unique and preferred DNA-binding sites of individual metals. We show how highly sensitive and selective DNA-based sensors can be designed that contain multiple binding sites for 21 heavy metal cations that bind to DNA and change its structure, consistent with the release of the DNA-bound dye.     

On the Stock Estimation for a Harvested Fish Population

We consider a stage-structured model of a harvested fish population and we are interested in the problem of estimating the unknown stock state for each class. The model used in this work to describe the dynamical evolution of the population is a discrete time system including a nonlinear recruitment relationship. To estimate the stock state, we build an observer for the considered fish model. This observer is an auxiliary dynamical system that uses the catch data over each time interval and gives a dynamical estimate of the stock state for each stage class. The observer works well even if the recruitment function in the considered model is not well known. The same problem for an age-structured model has been addressed in a previous work (Ngom et al., Math. Biosci. Eng. 5(2):337–354, 2008).