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Presented is a teaching progression that enhances learning through low-cost, manipulative transparencies. Discussed is instruction about restriction enzymes, plasmids, cutting plasmids, plasmid maps, recording data, and mapping restriction sites. Mapping wheels for student use is included. (CW)

Described is the role of technology in the Human Genome Project. A poster of the human genome as is known in 1990 is provided with a list of representative genes and a glossary of terms. (CW)

An activity that introduces students to the correct handling of bacterial recombinants, antibiotic sensitivity testing, insertional inactivation, plasmid DNA isolation, restriction endonuclease digestion, agarose gel electrophoresis, Southern blotting, hybridization, and autoradiography is presented. A list of needed materials, procedures, safety precautions, results, and discussion are included. (KR)

An experiment that uses the recombinant plasmid pX1108 to illustrate restriction mapping is described. The experiment involves three restriction enzymes and employs single and double restriction enzyme digestions. A list of needed materials, procedures, safety precautions, results, and discussion are included. (KR)

Describes the principal chemical substances which occur in most cells. These chemicals are the lipids, carbohydrates, proteins, and nucleic acids. Suggests that the structures of these substances be taught first since structure determines function. (JN)

Summarizes the current state of genetics, highlighting major historical events in the development of the field and discussing topics related to introns ("silent" or noncoding base sequences in eucaryotic genes) and exons (the coding parts of DNA). (JN)

Describes a senior-level course that: (1) focuses on the structure and reactions of macromolecules; (2) treats industrial polymers in a unified way; and (3) uses analysis of conformation and conformational statistics as a unifying approach. Also discusses course topics, including polysaccharides, proteins, nucleic acids, and others. (JN)

Examines what constraints govern the physical process of computation, considering such areas as whether a minimum amount of energy is required per logic step. Indicates that although there seems to be no minimum, answers to other questions are unresolved. Examples used include DNA/RNA, a Brownian clockwork turning machine, and others. (JN)

Explains the structural organization of DNA by providing information on the primary, secondary, tertiary, and higher organization levels of the molecule. Also includes illustrations and descriptions of sign-inversion and rotating models for supercoiling of DNA. (ML)

New advances in molecular biology have established a biotechnology industry and have changed ways people think about living things. In support of this theme, a discussion on historical development and current practice of gene cloning is presented. The role of nucleic acids, viruses, and therapeutic intervention is also considered. (DH)

The immune system includes the most diverse proteins known because they are encoded by hundreds of scattered gene fragments which can be combined in millions or billions of ways. Events of immune response, binding of antigens, antibody structure, T-cell receptors, and other immunologically-oriented topics are discussed. (DH)

Ribonucleic acid (RNA) converts genetic information into protein and usually must be processed to serve its function. RNA types, chemical structure, protein synthesis, translation, manufacture, and processing are discussed. Concludes that the first genes might have been spliced RNA and that humans might be closer than bacteria to primitive ancestors. (DH)

Structural form, bonding scheme, and chromatin structure of and gene-modification experiments with deoxyribonucleic acid (DNA) are described. Indicates that DNA's double helix is variable and also flexible as it interacts with regulatory and other molecules to transfer hereditary messages. (DH)

Reviews current findings that explain RNA's function as an enzyme in addition to being an informational molecule. Highlights recent research efforts and notes changes in the information base on RNA activity. Includes models and diagrams of RNA activity. (ML)

Discusses the applicability and advantages of using yeasts as popular and ideal model systems for studying and understanding eukaryotic biology at the cellular and molecular levels. Cites experimental tractability and the cooperative tradition of the research community of yeast biologists as reasons for this success. (RT)

Describes the initial site, and symptoms of the human immunodeficiency virus. Explains the diverse behavior and destructive consequences of the disease through discussion, diagrams, and pictures of the life cycle of the virus and the genetic material that controls it. (RT)

Presents insights into the endosymbiotic theory based on a re-examination of evidence from investigations into the archaebacteria and other strange organisms inhabiting the hindguts of wood-eating insects. Examines the mechanism of evolutionary change and the speed with which it occurs. (RT)

Examines the technical vocabulary of genetics as a source of error and confusion. Suggests that it is necessary to identify different types of problems associated with terminology and to organize them into logical classes to deal effectively with the difficulties. Highlights terms misused in textbooks. (RT)

Discussed is the Human Genome Project which aims to decipher the totality of the human genetic information. The historical background, the objectives, international cooperation, ethical discussion, and the role of UNESCO are included. (KR)

Because advanced students had difficulty in interpreting cleavage patterns obtained by gel electrophoresis related to rapid sequencing techniques for DNA and RNA, several formats were developed to aid in understanding this topic. Formats included print, print plus scrambled print, interactive computer-based instruction, and high-resolution computer graphics. (Author/JN)

Describes a unit on macromolecules that has been used in the 12th grade of many Israeli secondary schools. Topic areas in the unit include synthetic polymers, biological macromolecules, and nucleic acids. A unit outline is provided in an appendix. (JN)

Describes the use of a chart that systematically summarizes three basic steps that involve DNA and its decoding in both eukaryotic and prokaryotic cells: replication; transcription, and translation. Indicates that the chart (mounted on a tranparency) does an adequate job of conveying basic information about nucleic acids to students. (DH)

Describes advances made in the understanding of how sex hormones may modify various cognitive skills, how normal brain signaling mechanisms may cause nerve cell death, and how many cells appear to hold genetic agents which determine their own destruction. (RT)

Describes a way to help students grasp intermediate steps in the movement and relationships of the various components involved in the addition of an amino acid to a nascent peptide chain. Includes drawings of the model in operation, construction details, and suggested shapes and labeling of components. (RT)

In this article, background information on the photochemical properties of nucleic acids is provided and an experiment to illustrate this concept is presented. Sample preparation, procedures, and data analysis are discussed. Sample UV absorption spectra are included. (CW)

An imaging technique that uses fluorescent dyes and allows scientists to track DNA as it moves through gels or in solution is described. The importance, opportunities, and implications of this technique are discussed. (KR)

Discussed are novel ways of blocking gene expression by using antisense RNA. The use of antisense technology in plant development and its possible use in human disease therapy are described. (KR)

Discussed is a designer-drug technology called antisense which blocks messenger RNA's ability to carry information to protein producing sites in the cell. The applications of this drug to AIDS research, cancer therapy, and other diseases are discussed. (KR)

A single, didactic criterion for the description of all biological macromolecules is proposed. This criterion is applicable to globular, fibrous or mixed proteins, as well as to nucleic acids and lipids or polysaccharides. Specific examples are given for all except lipids and polysaccarides. (KR)

Provides sources of nonanimal biochemical materials (which are relatively inexpensive, readily available, and require no special storage) suitable for use in biochemistry experiments. They are presented under these headings: (1) enzymes and other proteins; (2) carbohydrates; (3) lipids; (4) nucleic acids; and (5) metabolism. (JN)

Described is a simple technique for isolating DNA from halophilic bacteria. Materials, procedure, and additional experiments are outlined. It is stated that the DNA obtained will be somewhat contaminated with cellular proteins and RNA. Offers a procedure for greater purification. (RT)

8-oxo-7,8-dihydro-2′-dexyoguanine (8-oxo-dG) can be tautomerized to a 6-enolate,8-keto tautomer through nearby-NH deprotonation at elevated pH. In this work, the N3-protonated 8-oxo-dG tautomers in deuterated pH-buffer solutions were studied using steady-state UV/Vis, FTIR, and ultrafast two-dimensional IR spectroscopies. The presence of 6,8-diketo and C6-anionic tautomers at neutral to basic conditions (pD = 7.4–12.0) was revealed by UV/Vis and FTIR results and was further confirmed by 2D IR signals in both diagonal and off-diagonal regions. However, the C6-enol tautomer, which may be an intermediate during the transition from 6,8-diketo to C6-enolate,C8-keto, was not observed appreciably due to its extreme low population. Furthermore, the neutral-to-anionic tautomeric transition of N3H-8-oxo-dG studied in this work occurs under more basic conditions than the N1H-8-oxo-dG reported previously, showing a higher pKa value for N3H than N1H. Finally, vibrational relaxation of the carbonyl stretching mode was found to be both molecular site dependent and pD dependent for 8oxo-dG. Taken together, this work shows that the ultrafast infrared spectroscopic method is effective for examining tautomers and their dynamics in nucleic acids. [ABSTRACT FROM AUTHOR]

The folding of RNA and DNA strands plays crucial roles in biological systems and bionanotechnology. However, studying these processes with high-resolution numerical models is beyond current computational capabilities due to the timescales and system sizes involved. In this article, we present a new coarse-grained model for investigating the folding dynamics of nucleic acids. Our model represents three nucleotides with a patchy particle and is parameterized using well-established nearest-neighbor models. Thanks to the reduction of degrees of freedom and to a bond-swapping mechanism, our model allows for simulations at timescales and length scales that are currently inaccessible to more detailed models. To validate the performance of our model, we conducted extensive simulations of various systems: We examined the thermodynamics of DNA hairpins, capturing their stability and structural transitions, the folding of an MMTV pseudoknot, which is a complex RNA structure involved in viral replication, and also explored the folding of an RNA tile containing a k-type pseudoknot. Finally, we evaluated the performance of the new model in reproducing the melting temperatures of oligomers and the dependence on the toehold length of the displacement rate in toehold-mediated displacement processes, a key reaction used in molecular computing. All in all, the successful reproduction of experimental data and favorable comparisons with existing coarse-grained models validate the effectiveness of the new model. [ABSTRACT FROM AUTHOR]

The accurate detection and quantification of biological species that are rarely present but potentially devastating is of paramount importance for the life sciences, biosecurity, food safety, and environmental monitoring. Consequently, there has been significant interest in the sensitive and accurate detection of nucleic acids, leveraging both chemical and biological methods. Among these, quantitative polymerase chain reaction (qPCR) is regarded as the gold standard due to its sensitivity and precision in identifying specific nucleic acid targets. Despite the widespread adoption of qPCR for nucleic acid detection, the analysis of qPCR data typically depends on the use of calibrated standard curves and a threshold method to interpret signal measurements. In this study, we use a stochastic simulation to show the limitations of the threshold method due to its assumptions on amplification kinetics. We propose a new approach for the absolute quantification of nucleic acids that overcomes these limitations by reconstructing the efficiency profile across amplification cycles and using cumulative amplification folds to build a standard curve, thus avoiding the constant efficiency assumption. Our method, validated through experiments with nucleic acid amplification in the presence of potent inhibitors, demonstrates improved accuracy in quantifying nucleic acids, avoiding the systematic errors of the threshold method. This innovation enhances the reliability of nucleic acid quantification, especially where traditional methods struggle with kinetic variability. [ABSTRACT FROM AUTHOR]

We herein present a multifunctional self-priming hairpin probe-based isothermal amplification, termed MSH, enabling one-pot detection of target nucleic acids. The sophisticatedly designed multifunctional self-priming hairpin (MSH) probe recognizes the target and rearranges to prime itself, triggering the amplification reaction powered by the continuously repeated extension, nicking, and target recycling. As a consequence, a large number of double-stranded DNA (dsDNA) amplicons are produced that could be monitored in real-time using a dsDNA-intercalating dye. Based on this unique design approach, the nucleocapsid (N) and the open reading frame 1 ab (ORF1ab) genes of SARS-CoV-2 were successfully detected down to 1.664 fM and 0.770 fM, respectively. The practical applicability of our method was validated by accurately diagnosing 60 clinical samples with 93.33% sensitivity and 96.67% specificity. This isothermal one-pot MSH technique holds great promise as a point-of-care testing protocol for the reliable detection of a wide spectrum of pathogens, particularly in resource-limited settings.

Theodor (Ted) Otto Diener (* 28 February 1921 in Zürich, Switzerland; † 28 March 2023 in Beltsville, MD, USA) pioneered research on viroids while working at the Plant Virology Laboratory, Agricultural Research Service, USDA, in Beltsville. He coined the name viroid and defined viroids important features like the infectivity of naked single-stranded RNA without protein-coding capacity. During scientific meetings in the 1970s and 1980s, viroids were often discussed at conferences together with other subviral pathogens. This term includes what are now called satellite RNAs and prions. Satellite RNAs depend on a helper virus and have linear or, in the case of virusoids, circular RNA genomes. Prions, proteinaceous infectious particles, are the agents of scrapie, kuru and some other diseases. Many satellite RNAs, like viroids, are non-coding and exert their function by thermodynamically or kinetically controlled folding, while prions are solely host-encoded proteins that cause disease by misfolding, aggregation and transmission of their conformations into infectious prion isoforms. In this memorial, we will recall the work of Ted Diener on subviral pathogens.

Silver cations can mediate base pairing of guanine (G) DNA oligomers, yielding linear parallel G-Ag+-G duplexes with enhanced stabilities compared to those of canonical DNA duplexes. To enable their use in programmable DNA nanotechnologies, it is critical to understand solution-state formation and the nanomechanical stiffness of G-Ag+-G duplexes. Using temperature-controlled circular dichroism (CD) spectroscopy, we find that heating mixtures of G oligomers and silver salt above 50 °C fully destabilizes G-quadruplex structures and converts oligomers to G-Ag+-G duplexes. Electrospray ionization mass spectrometry supports that G-Ag+-G duplexes form at stoichiometries of 1 Ag+ per base pair, and CD spectroscopy suggests that as the Ag+/base stoichiometry increases further, G-Ag+-G duplexes undergo additional morphological changes. Using liquid-phase atomic force microscopy, we find that this excess Ag+ enables assembly of long fiberlike structures with ∼2.5 nm heights equivalent to a single DNA duplex but with lengths that far exceed a single duplex. Finally, using the conditions established to form single G-Ag+-G duplexes, we use a surface forces apparatus (SFA) to compare the solution-phase stiffness of single G-Ag+-G duplexes with dG-dC Watson-Crick-Franklin duplexes. SFA shows that G-Ag+-G duplexes are 1.3 times stiffer than dG-dC duplexes, confirming gas-phase ion mobility spectrometry measurements and computational predictions. These findings may guide the development of structural DNA nanotechnologies that rely on silver-mediated base pairing.

Regulation of protein mobility is a fundamental aspect of cellular processes. In this study, we examined the impact of DNA methylation on the diffusion of nucleoid associated protein Hfq. This protein is one of the most abundant proteins that shapes the bacterial chromosome and is involved in several aspects of nucleic acid metabolism. Fluorescence microscopy was employed to monitor the movement of Hfq along doublestranded DNA, which was stretched due to confinement within a nanofluidic channel. The mobility of Hfq is significantly influenced by DNA methylation. Our results underscore the importance of bacterial epigenetic modifications in governing the movement of nucleoid associated proteins such as Hfq. Increased levels of methylation result in enhanced binding affinity, which in turn slows down the diffusion of Hfq on DNA. The reported control of protein mobility by DNA methylation has potential implications for the mechanisms involved in target DNA search processes and dynamic modelling of the bacterial chromosome. [ABSTRACT FROM AUTHOR]

In this study, a novel electrochemical biosensor was developed for the sensitive and selective detection of the Acinetobacter baumannii gene sequence. The biosensor was created by immobilizing a capture probe specific to the A. baumannii gene on the surface of chitosan-gold modified pencil graphite electrodes. Following solid-state hybridization on the Chit-Au/PGE surface, the target DNA sequence of the A. baumannii was detected by measuring the guanine signal using square wave voltammetry (SWV). All experimental parameters impacting sensor response are examined in order to improve hybridization efficacy, and the electrochemical biosensor's performance. The limit of detection (LOD) for the A. baumannii gene sequence was calculated and found to be 1.93 nM. Three different non-complementary DNA sequences were used to evaluate the assay selectivity, but no interference effect was obtained. Additionally, the potential applicability of the biosensor to real samples was tested in artificial serum media. The suggested electrochemical test procedure is simple, approachable, and quick, making it a convenient approach for the screening of DNA sequence. [Display omitted] • Pencil graphite electrodes were modified with chitosan-Au composite. • CHIT-Au/PGEs were used for electrochemical biosensing of A. baumannii gene • The [Fe(CN) 6 ]3-/4- oxidation signal was measured by SWV technique. • The proposed electrochemical assay is simple, rapid, and accessible. [ABSTRACT FROM AUTHOR]

Cells use transient membraneless organelles to regulate biological reaction networks. For example, stress granules selectively store mRNA to downregulate protein expression in response to heat or oxidative stress. Models mimicking this active behavior should be established to better understand in vivo regulation involving compartmentalization. Here we use active, complex coacervate droplets as a model for membraneless organelles to spatiotemporally control the activity of a catalytic DNA (DNAzyme). Upon partitioning into these peptide‐RNA droplets, the DNAzyme unfolds and loses its ability to catalyze the cleavage of a nucleic acid strand. We can transiently pause the DNAzyme activity upon inducing droplet formation with fuel. After fuel consumption, the DNAzyme activity autonomously restarts. We envision this system could be used to up and downregulate multiple reactions in a network, helping understand the complexity of a cell's pathways. By creating a network where the DNAzyme could reciprocally regulate the droplet properties, we would have a powerful tool for engineering synthetic cells. [ABSTRACT FROM AUTHOR]

Pharyngitis is an inflammatory condition of the pharynx and/or tonsils commonly seen in both children and adults. Viruses and bacteria represent the most common encountered etiologic agents—yeast/fungi and parasites are infrequently implicated. Some of these are predominantly observed in unique populations (eg, immunocompromised or unvaccinated individuals). This manuscript (part 2 of 3) summarizes the current state of laboratory and point-of-care diagnostic testing and highlights the expanding role of nucleic acid amplification in the expedited diagnosis and management of patients with acute pharyngitis. It discusses preanalytical, analytical, and postanalytical variables that impact the performance of culture, rapid antigen, and nucleic acid amplification testing. Finally, it sets the stage for part 3, which discusses the emerging role of biomarkers in the management of individuals with acute pharyngitis. [ABSTRACT FROM AUTHOR]

The appraisal of drug–surfactant binding interactions is pharmacologically and pharmaceutically noteworthy. The simple model compounds of nucleic acids such as nucleic acid bases/nucleosides are key biologically active molecules useful for studying interactions of nucleic acids with other components in bio-fluids. Therefore, the binding studies of promethazine hydrochloride (PMT) drug with individual surfactants such as sodium dodecylsulphate (SDS, anionic surfactant), sodium lauroylsarcosinate (SLS, amino based-surfactant), and sodium cholate (NaC, trihydroxy bile salt), and their mixed systems (PMT + SDS/SLS/NaC) at mole fractions, αPMT = 0.02, 0.04, 0.06, and 0.08 have been accomplished in water and 0.025 mol kg−1 aqueous uridine (nucleoside) solutions at temperatures (298.15, 308.15, and 318.15) K. Thermodynamic characteristics of micellization have been obtained from the conductivity studies. Binding constants (Kb, Kf) and standard Gibbs free energies of binding (ΔGοb, ΔGοf) for pure PMT drug and its mixed systems with anionic surfactants were evaluated from UV–visible absorption and fluorescence emission studies. Further, the density functional theory (DFT) calculations for studied systems were carried out to get net stabilization energies for the drug–surfactant complexes. NBO and HOMO–LUMO analyses were also carried out in the absence/presence of uridine. Dynamic light scattering study was used to measure the hydrodynamic diameters in micellar systems. The major influences of hydrophobic–hydrophobic interactions among the hydrophobic groups and electrostatic (ionic and H-bond) interactions among the hydrophilic/ionic groups of PMT drug, SDS/SLS/NaC, and uridine have been depicted from the present investigations. [ABSTRACT FROM AUTHOR]

Inositol pyrophosphates (PP-InsPs) are nutrient messengers whose cellular levels are precisely regulated. Diphosphoinositol pentakisphosphate kinases (PPIP5Ks) generate the active signaling molecule 1,5-InsP8. PPIP5Ks harbor phosphatase domains that hydrolyze PP-InsPs. Plant and Fungi Atypical Dual Specificity Phosphatases (PFA-DSPs) and NUDIX phosphatases (NUDTs) are also involved in PP-InsP degradation. Here, we analyze the relative contributions of the three different phosphatase families to plant PP-InsP catabolism. We report the biochemical characterization of inositol pyrophosphate phosphatases from Arabidopsis and Marchantia polymorpha. Overexpression of different PFA-DSP and NUDT enzymes affects PP-InsP levels and leads to stunted growth phenotypes in Arabidopsis. nudt17/18/21 knock-out mutants have altered PP-InsP pools and gene expression patterns, but no apparent growth defects. In contrast, Marchantia polymorpha Mppfa-dsp1ge, Mpnudt1ge and Mpvip1ge mutants display severe growth and developmental phenotypes and associated changes in cellular PP-InsP levels. Analysis of Mppfa-dsp1geand Mpvip1ge mutants supports a role for PP-InsPs in Marchantia phosphate signaling, and additional functions in nitrate homeostasis and cell wall biogenesis. Simultaneous elimination of two phosphatase activities enhanced the observed growth phenotypes. Taken together, PPIP5K, PFA-DSP and NUDT inositol pyrophosphate phosphatases regulate growth and development by collectively shaping plant PP-InsP pools. Author summary: Organisms must maintain adequate levels of nutrients in their cells and tissues. One such nutrient is phosphorus, an essential building block of cell membranes, nucleic acids and energy metabolites. Plants take up phosphorus in the form of inorganic phosphate and require adequate phosphate levels to support their growth and development. It has been shown that plants and other eukaryotic organisms "measure" cellular phosphate levels using inositol pyrophosphate signaling molecules. The concentration of inositol pyrophosphates serves as a proxy for the cellular concentration of inorganic phosphate, and therefore inositol pyrophosphate synthesis and degradation must be tightly regulated. Here, we report that three different families of enzymes contribute to the degradation of inositol pyrophosphates in plants. Taken together, the different phosphatases shape cellular inositol pyrophosphate pools to regulate inorganic phosphate levels. Loss-of-function mutants of the different enzymes show altered nitrate levels and changes in cell wall architecture, suggesting that inositol pyrophosphates may regulate cellular processes beyond phosphate homeostasis. [ABSTRACT FROM AUTHOR]

Opsonization of red blood cells that retain mitochondria (Mito+ RBCs), a feature of systemic lupus erythematosus (SLE), triggers type I interferon (IFN) production in macrophages. We report that monocytes (Mos) co-produce IFN and mature interleukin-1β (mIL-1β) upon Mito+ RBC opsonization. IFN expression depended on cyclic GMP-AMP synthase (cGAS) and RIG-I-like receptors' (RLRs) sensing of Mito+ RBC-derived mitochondrial DNA (mtDNA) and mtRNA, respectively. Interleukin-1β (IL-1β) production was initiated by the RLR antiviral signaling adaptor (MAVS) pathway recognition of Mito+ RBC-derived mtRNA. This led to the cytosolic release of Mo mtDNA, which activated the inflammasome. Importantly, mIL-1β secretion was independent of gasdermin D (GSDMD) and pyroptosis but relied on IFN-inducible myxovirus-resistant protein 1 (MxA), which facilitated the incorporation of mIL-1β into a trans -Golgi network (TGN)-mediated secretory pathway. RBC internalization identified a subset of blood Mo expressing IFN-stimulated genes (ISGs) that released mIL-1β and expanded in SLE patients with active disease. [Display omitted] • Mos co-produce type I IFN and IL-1β upon Mito+ RBC opsonization • IFN and IL-1β expression rely on sensing RBC-derived mtDNA and mtRNA • IL-1β secretion is MxA mediated and independent of GSDMD and pyroptosis • GPA+ blood Mos expressing IFN and IL-1β are expanded in active SLE SLE is an interferonopathy, but blocking IFN does not benefit all patients. Caielli et al. found that SLE monocytes co-produce IFN and IL-1β, a phenotype recapitulated upon in vitro Mito+ RBC opsonization. Cytosolic sensing of mitochondrial nucleic acids induces MxA, which enables IL-1β secretion independently of GSDMD and pyroptosis. [ABSTRACT FROM AUTHOR]