Application Of DNA Microarrays Research Articles

DNA Microarray Technology Principles and Applications in Genetic Research

DNA microarray technology, a high-throughput gene analysis tool, has played a crucial role in genetic research. This paper explores the fundamental principles of DNA microarray technology, including its working mechanisms, sample preparation, and types of chips. It emphasizes the applications of DNA microarrays in detecting genetic variations and analyzing gene expression profiles, illustrating examples in single nucleotide polymorphism (SNP) analysis and genome-wide association studies (GWAS). The paper also discusses the technical details of high-throughput gene expression measurement and differential expression analysis. Additionally, it analyzes the main advantages of DNA microarray technology, such as high throughput and sensitivity, while also noting its limitations, including data interpretation complexity and technical costs. Finally, the paper looks forward to future development trends in DNA microarray technology, particularly its potential applications in personalized medicine.

Correlated Hybrid DNA Structures Explored by the oxDNA Model.

Thermodynamically, perfect DNA hybridization can be formed between probes and their corresponding targets due to the favorable energy. However, this is not the case dynamically. Here, we use molecular dynamics (MD) simulations based on the oxDNA model to investigate the process of DNA microarray hybridization. In general, correlated hybrid DNA structures are formed, including one probe associated with several targets as well as one target hybrid with multiple probes leading to the target-mediated hybridization. The formation of these two types of correlated structures largely depends on the surface coverage of the DNA microarray. Moreover, DNA sequence, DNA length, and spacer length have an impact on the structural formation. Our findings shed light on the dynamics of DNA hybridization, which is important for the application of DNA microarray.

Hydrolysis of Oligodeoxyribonucleotides on the Microarray Surface and in Solution by Catalytic Anti-DNA Antibodies in Systemic Lupus Erythematosus.

Anti-DNA antibodies are known to be classical serological hallmarks of systemic lupus erythematosus (SLE). In addition to high-affinity antibodies, the autoantibody pool also contains natural catalytic anti-DNA antibodies that recognize and hydrolyze DNA. However, the specificity of such antibodies is uncertain. In addition, DNA binding to a surface such as the cell membrane, can also affect its recognition by antibodies. Here, we analyzed the hydrolysis of short oligodeoxyribonucleotides (ODNs) immobilized on the microarray surface and in solution by catalytic anti-DNA antibodies from SLE patients. It has been shown that IgG antibodies from SLE patients hydrolyze ODNs more effectively both in solution and on the surface, compared to IgG from healthy individuals. The data obtained indicate a more efficient hydrolysis of ODNs in solution than immobilized ODNs on the surface. In addition, differences in the specificity of recognition and hydrolysis of certain ODNs by anti-DNA antibodies were revealed, indicating the formation of autoantibodies to specific DNA motifs in SLE. The data obtained expand our understanding of the role of anti-DNA antibodies in SLE. Differences in the recognition and hydrolysis of surface-tethered and dissolved ODNs need to be considered in DNA microarray applications.

Recent Progress in Development and Application of DNA, Protein, Peptide, Glycan, Antibody, and Aptamer Microarrays.

Microarrays are one of the trailblazing technologies of the last two decades and have displayed their importance in all the associated fields of biology. They are widely explored to screen, identify, and gain insights on the characteristics traits of biomolecules (individually or in complex solutions). A wide variety of biomolecule-based microarrays (DNA microarrays, protein microarrays, glycan microarrays, antibody microarrays, peptide microarrays, and aptamer microarrays) are either commercially available or fabricated in-house by researchers to explore diverse substrates, surface coating, immobilization techniques, and detection strategies. The aim of this review is to explore the development of biomolecule-based microarray applications since 2018 onwards. Here, we have covered a different array of printing strategies, substrate surface modification, biomolecule immobilization strategies, detection techniques, and biomolecule-based microarray applications. The period of 2018-2022 focused on using biomolecule-based microarrays for the identification of biomarkers, detection of viruses, differentiation of multiple pathogens, etc. A few potential future applications of microarrays could be for personalized medicine, vaccine candidate screening, toxin screening, pathogen identification, and posttranslational modifications.

Optimization of Functional Group Concentration of N, N-Dimethylacrylamide-based Polymeric Coatings and Probe Immobilization for DNA and Protein Microarray Applications

We report here a deep investigation into the effect of the concentration of a polymeric coating's functional groups on probe density immobilization with the aim of establishing the optimal formulation to be implemented in specific microarray applications. It is widely known that the ideal performance of a microarray strictly depends on the way probes are tethered to the surface since it influences the way they interact with the complementary target. The N, N-dimethylacrylamide-based polymeric coating introduced by our research group in 2004 has already proven to offer great flexibility for the customization of surface properties; here, we demonstrate that it also represents the perfect scaffold for the modulation of probe grafting. With this aim in mind, polymers with increasing concentrations of N-acryloyloxysuccinimide (NAS) were synthesized and the coating procedure optimized accordingly. These were then tested not only in DNA microarray assays, but also using protein probes (with different MWs) to establish which formulation improves the assay performance in specific applications. The flexibility of this polymeric platform allowed us also to investigate a different immobilization chemistry-specifically, click chemistry reactions, thanks to the insertion of azide groups into the polymer chains-and to evaluate possible differences generated by this modification.

Prospects of DNA microarray application in management of chronic obstructive pulmonary disease: A systematic review

Abstract Chronic obstructive pulmonary disease (COPD) is incurable chronic disease which kills 3.3 million each year worldwide. Number of global cases of COPD is steadily rising alongside with life expectancy, disproportionally hitting middle-income countries like Russia and China, in such conditions, new approaches to the COPD management are desperately needed. DNA microarray technology is a powerful genomic tool that has the potential to uncover underlying COPD biological alteration and brings up revolutionized treatment option to clinicians. We executed systematic review studies of studies published in last 10 years regarding DNA microarray application in COPD management, with complacence to PRISMA criteria and using PubMed and Medline data bases as data source. Out of 920 identified papers, 39 were included in the final analysis. We concluded that Genome-wide expression profiling using DNA microarray technology has great potential in enhancing COPD management. Current studied proofed this method is reliable and possesses many potential applications such as individual at risk of COPD development recognition, early diagnosis of disease, COPD phenotype identification, exacerbation prediction, personalized treatment optioning and prospect of oncogenesis evaluation in patients with COPD. Despite all the proofed benefits of this technology, researchers are still in the early stage of exploring it's potential. Therefore, large clinical trials are still needed to set up standard for DNA microarray techniques usage implementation in COPD management guidelines, subsequently giving opportunity to clinicians for controlling or even eliminating COPD entirely.

Application of DNA Microarray in Genetic Mutation Detection in Patients with Thalassemia

To perform dried blood spots thalassemia gene detection in patients with positive blood phenotypes by microarray technology, and evaluate its value in clinical detection. DNA samples were extracted from dried blood spots of 410 patients. Microarray technology was used to detect 3 deletion and 3 non-deletion types of α-thalassemia and 19 β-thalassemia point mutations which were common gene mutions in China. There were 357 positive cases in all the 410 tested samples with the positive rate 87.07%, among which 299 cases (72.93%) carried deletion or point mutations of α-thalassemia, 29 cases (7.07%) carried point mutations of β-thalassemia and 29 cases (7.07%) carried gene mutations of complex αβ-thalassemia syndrome. The mutations of α-thalassemia were involved with --SEA heterozygous deletion (177 cases, 59.2%), αCS heterozygote (60 cases, 20.07%) and several other genotypes. The common mutations of β- thalassemia were involved with βCD41-42 heterozygote (10 cases, 34.48%) and βCD17 heterozygote (9 cases, 31.03%). The mutations of complex αβ-thalassemia syndrome were mainly involved with --SEA/αα+βCD17/βN (7 cases, 24.14%), αCSα/αα + βCD41-42/βN (3 cases, 10.34%) and -α4.2/αα + βCD17/βN (3 cases, 10.34%). The most common genetic mutations are --SEA for α-thalassemia and CD41-42 for β-thalassemia in Liuzhou, Guangxi Zhuang Autonomous Region. A and β-thalassemia can be detected at the same time by microarray chip technology in a high throughput manner.

Profiling gene expression dynamics underpinning conventional testing approaches to better inform pre-clinical evaluation of an age appropriate spironolactone formulation

There is a need to accelerate paediatric formulation evaluation and enhance quality of early stage data in drug development to alleviate the information pinch point present between formulation development and clinical evaluation. This present work reports application of DNA microarrays as a high throughput screening tool identifying markers for prediction of bioavailability and formulation driven physiological responses. With a focus on enhancing paediatric medicine provision, an oral liquid spironolactone suspension was formulated addressing a paediatric target product profile. Caco-2 cells cultured on transwell inserts were implemented in transport assays in vitro and DNA microarrays were used to examine gene expression modulation. Wistar rats were used to derive in vivo bioavailability data. In vitro, genomic, and in vivo data sets were concurrently evaluated linking drug transport and the genomic fingerprint generated by spironolactone formulation exposure. Significant changes in gene expression are reported as a result of formulation exposure. These include genes coding for ATP-binding cassette (ABC) transporters, solute carrier (SLC) transporters, cytochrome P450 (CYP) enzymes, and carboxylesterase enzymes. Genomic findings better inform pre-clinical understanding of pharmacokinetic and pharmacodynamic responses to spironolactone and its active metabolites than current in vitro drug transport assays alone.

A Conversation with Grace Wahba

Grace Wahba (née Goldsmith, born August 3, 1934), I. J. Schoenberg-Hilldale Professor of Statistics at the University of Wisconsin-Madison (Emerita), is a pioneer in methods for smoothing noisy data. Her research combines theoretical analysis, computation and methodology motivated by innovative scientific applications. Best known for the development of generalized cross-validation (GCV), the connection between splines and Bayesian posterior estimates, and “Wahba’s problem,” she has developed methods with applications in demographic studies, machine learning, DNA microarrays, risk modeling, medical imaging and climate prediction. Grace grew up in the Washington, DC area and New Jersey, and graduated from Montclair High School. She was educated at Cornell (B.A. 1956), University of Maryland, College Park (M.A. 1962) and Stanford (Ph.D. 1966), and worked in industry for several years before receiving her doctorate in 1966 and settling in Madison in 1967. Although holding several visiting appointments, she has made Madison her home for over 50 years. She is the author of Spline Models for Observational Data which has garnered more than 8000 citations. Grace is treasured as an academic advisor and has mentored 39 Ph.D. students that have resulted in more than 330 academic descendants. She was elected to the United States National Academy of Sciences in 2000 and received an honorary degree of Doctor of Science from the University of Chicago in 2007. Wahba is a Fellow of several academic societies including the American Academy of Arts and Sciences, the American Association for the Advancement of Science, the American Statistical Association and the Institute of Mathematical Statistics. Over the years, she has received a selection of notable awards in the statistics community: R. A. Fisher Lectureship (2014), Gottfried E. Noether Senior Researcher Award (2009), Committee of Presidents of Statistical Societies Elizabeth Scott Award (1996) and the first Emanuel and Carol Parzen Prize for Statistical Innovation (1994).

Digital DNA microarray generation on glass substrates

In this work we show how DNA microarrays can be produced batch wise on standard microscope slides in a fast, easy, reliable and cost-efficient way. Contrary to classical microarray generation, the microarrays are generated via digital solid phase PCR. We have developed a cavity-chip system made of a PDMS/aluminum composite which allows such a solid phase PCR in a scalable and easy to handle manner. For the proof of concept, a DNA pool composed of two different DNA species was used to show that digital PCR is possible in our chips. In addition, we demonstrate that DNA microarray generation can be realized with different laboratory equipment (slide cycler, manually in water baths and with an automated cartridge system). We generated multiple microarrays and analyzed over 13,000 different monoclonal DNA spots to show that there is no significant difference between the used equipment. To show the scalability of our system we also varied the size and number of the cavities located in the array region up to more than 30,000 cavities with a volume of less than 60 pL per cavity. With this method, we present a revolutionary tool for novel DNA microarrays. Together with new established label-free measurement systems, our technology has the potential to give DNA microarray applications a new boost.

The preparation and clinical application of diagnostic DNA microarray for the detection of pathogens in intracranial bacterial and fungal infections

The present study prepared 2 types of DNA diagnostic chips based on 16S ribosomal DNA (rDNA) and 18S-28S rDNA, and evaluated their values in the detection of pathogens in intracranial bacterial/fungal infections. A total of 14 probes of bacteria (Klebsiella pneumonia, Acinetobacter baumannii, Pseudomonas aeruginosa, Escherichia coli, Haemophilus influenza, Stenotrophomonas maltophilia, Neisseria meningitidis, Enterobacter spp., Enterococcus faecalis, Enterococcus faecium, Listeria monocytogenes, Staphylococcus aureus, Streptococcus pneumonia and coagulase negative staphylococcus) and 4 probes of fungi (Candida albicans, Candida tropicalis, Candida glabrata and Cryptococcus neoformans), determined frequently in cerebrospinal fluid (CSF), were designed and used for preparation of microarrays. CSF samples from 88 patients with clinically suspected intracranial infection and standard strains were used to evaluate the chips. The same samples were also analyzed by culture and sequencing. The results demonstrated that the sensitivity, specificity and false-positive rate of the microarray assay compared with culture method were 100 vs. 68.3% (P<0.05), 97.1 vs. 100%, and 2.9 vs. 0%, respectively. The minimum concentration of detection with the chips was 10 cfu ml−1 for bacteria and 100 cfu ml−1 for fungi. The specificity of the probes was confirmed, and no cross-reaction was detected in the present study. Furthermore, 13 cases were positive in the microarray and negative in culture. However, 4 cases were not identified as clear pathogens and only positive in the 16S probe sites. The diagnostic DNA microarray for intracranial infections has proven to be more rapid and sensitive, and it may be a better option for clinical application than culture methods.

Application of DNA Microarrays in Cytogenetics

Application of DNA Microarrays in Cytogenetics

Optimization of Cyanine Dye Stability and Analysis of FRET Interaction on DNA Microarrays

The application of DNA microarrays for high throughput analysis of genetic regulation is often limited by the fluorophores used as markers. The implementation of multi-scan techniques is limited by the fluorophores’ susceptibility to photobleaching when exposed to the scanner laser light. This paper presents combined mechanical and chemical strategies which enhance the photostability of cyanine 3 and cyanine 5 as part of solid state DNA microarrays. These strategies are based on scanning the microarrays while the hybridized DNA is still in an aqueous solution with the presence of a reductive/oxidative system (ROXS). Furthermore, the experimental setup allows for the analysis and eventual normalization of Förster-resonance-energy-transfer (FRET) interaction of cyanine-3/cyanine-5 dye combinations on the microarray. These findings constitute a step towards standardization of microarray experiments and analysis and may help to increase the comparability of microarray experiment results between labs.

Kinetic discrimination of DNA single-base mutations by localized surface plasmon resonance

Clinical application of DNA microarrays used for screening of single nucleotide polymorphisms (SNPs) are very important for diagnosis of diseases and appropriate treatment of patients. In this paper localized surface plasmon resonance (LSPR) technique has been used to study the DNA hybridization process for binary solutions of respectively perfectly matching (PM) and single base mismatching (MM) 93-mer ssDNA from KRAS codon 12. 5′-thiol modified 35-mer ssDNA has been linked to the Au nanodisks array as probe with a surface coverage of 2.8±0.1×1012/cm2. Probe’s binding properties was investigated in details, obtaining a sensitivity down to 10nMand 13nM, respectively for PM and MM, showing that the hybridization process occurs at a lower rate for MM with respect to PM target. The competitive hybridization is accounted for by an inhibition model, where the non-complementary sequences kinetically hinder the hybridization of the perfect matching sequences, owing to their above mentioned affinity constant differences for the same probe. Accordingly, the single nucleotide polymorphisms can therefore be revealed in a single step and label free mode with high sensitivity and specificity by LSPR measurements.

Semidefinite Programming for Computable Performance Bounds on Block-Sparsity Recovery

In this paper, we use fixed point theory and semidefinite programming to compute the performance bounds on convex block-sparsity recovery algorithms. As a prerequisite for optimal sensing matrix design, computable performance bounds would open doors for wide applications in sensor arrays, radar, DNA microarrays, and many other areas where block-sparsity arises naturally. We define a family of quality measures for arbitrary sensing matrices as the optimal values of certain optimization problems. The reconstruction errors of convex recovery algorithms are bounded in terms of these quality measures. We demonstrate that as long as the number of measurements is relatively large, these quality measures are bounded away from zero for a large class of random sensing matrices, a result parallel to the probabilistic analysis of the block restricted isometry property. As the primary contribution of this work, we associate the quality measures with the fixed points of functions defined by a series of semidefinite programs. This relation with fixed point theory yields polynomial-time algorithms with global convergence guarantees to compute the quality measures.

Differential Hepatic Gene Expression Profile and Histone Modification After Acute or Chronic Hyperthyroidism or During Remission.

In acute and chronic hyperthyroid patients, clinical symptoms may not always reflect their thyroid hormone status. After normalization of serum thyroid hormone concentrations in these patients, some symptoms caused by hyperthyroidism may remain unchanged (1). Even in subclinical hyperthyroid patients, functional recovery after restoration to euthyroidism is sometimes incomplete (2). The underlying mechanisms of such incomplete recovery are not yet fully understood. Indeed, functional adaptation or irreversible alteration may be induced during exposure to excess thyroid hormone. The liver is a representative target organ of thyroid hormone action, where T3 regulates various hepatic functions including cholesterol, triglyceride, and carbohydrate metabolism (3). To accomplish this, T3 regulates the expression of many rate-limiting enzymes of such metabolic pathways. In addition, thyroid hormone signaling activates autophagy, which plays a key role in regulating lipid metabolism, mitochondrial quality, and various proteins regulating metabolic pathways in hepatocytes (4). Thus, dysregulation of thyroid hormone action in liver may induce serious clinical conditions such as nonalcoholic fatty liver disease (5). To examine the role of thyroid hormone on the expression of hepatic genes, several groups have applied DNA microarray analysis to the hepatic gene expression profile (6). In these studies, it has become clear that the transcriptomic response is different between acute and chronic T3 treatment (7). However, the expression profile of T3-sensitive genes after amelioration of hyperthyroidism has not yet been studied. In this issue of Endocrinology, Ohba et al (8) examined the effects of a single T3 injection, chronic T3 injections (14 d), or chronic injections followed T3 withdrawal for 10 days on the hepatic gene expression profile of adult male mice using gene expression microarrays. Because they used a large dose of T3 (20 g/100 g body weight), all mice became hyperthyroid, which was confirmed by suppressed TSH levels. TSH levels then increased transiently 5 days after the last T3 injection, before normalizing by 10 days. They identified 680 genes that were induced by either a single or chronic injection of T3. Interestingly, only 10% of these genes were induced by both treatments, indicating that most acutely induced T3-sensitive genes became desensitized to the administration dose over time. Furthermore, 10% of genes did not return to normal levels of expression despite normalization of thyroid hormone concentrations after T3 withdrawal. Although Ohba et al (8) found many genes that were altered by T3 treatment or its withdrawal, such genes may not be directly regulated by thyroid hormone. Thus, they studied further the expression of 8 genes (B cell lymphoma 3 [Bcl3], cytochrome P450 17 -hydroxylase [Cyp17a1], Cyp 17,20-lyase [Cpt1a], fatty acid synthase [Fasn], fibroblast growth factor 21 [Fgf21], isocitrate dehydrogenase 3 [Idh3a], thyroid hormone-responsive protein [Thrsp], and type-1 iodothyronine deiodinase [Dio1]) thought to have functional thyroid hormone response elements. Interestingly, despite this, the expression pattern was not always the same. Among 6 genes that were induced by acute T3 treatment (Bcl3, Cpt1a, Dio1, Fasn, Idh3a, and Thrsp), the expression of 4 genes (Bcl3, Cpt1a, Fasn, and Thrsp) was not elevated after 14 days of T3 treatment, indicating desensitization by chronic treatment, whereas both Dio1 and Idh3a were induced by both

Application of DNA Microarray to Clinical Diagnostics.

Microarray-based technology to conduct array comparative genomic hybridization (aCGH) has made a significant impact on the diagnosis of human genetic diseases. Such diagnoses, previously undetectable by traditional G-banding chromosome analysis, are now achieved by identifying genomic copy number variants (CNVs) using the microarray. Not only can hundreds of well-characterized genetic syndromes be detected in a single assay, but new genomic disorders and disease-causing genes can also be discovered through the utilization of aCGH technology. Although other platforms such as single nucleotide polymorphism (SNP) arrays can be used for detecting CNVs, in this chapter we focus on describing the methods for performing aCGH using Agilent oligonucleotide arrays for both prenatal (e.g., amniotic fluid and chorionic villus sample) and postnatal samples (e.g., blood).

SiPM as miniaturised optical biosensor for DNA-microarray applications

A miniaturized optical biosensor for low-level fluorescence emitted by DNA strands labelled with CY5 is showed. Aim of this work is to demonstrate that a Si-based photodetector, having a low noise and a high sensitivity, can replace traditional detection systems in DNA-microarray applications. The photodetector used is a photomultiplier (SiPM), with 25pixels. It exhibits a higher sensitivity than commercial optical readers and we experimentally found a detection limit for spotted dried samples of ∼1nM. We measured the fluorescence signal in different operating conditions (angle of analysis, fluorophores concentrations, solution volumes and support). Once fixed the angle of analysis, for samples spotted on Al-TEOS slide dried, the system is proportional to the concentration of the analyte in the sample and is linear in the range 1nM–1μM. For solutions, the range of linearity ranges from 100fM to 10nM. The system potentialities and the device low costs suggest it as basic component for the design and fabrication of a cheap, easy and portable optical system.

Age-associated murine cardiac lesions are attenuated by the mitochondria-targeted antioxidant SkQ1.

Age-related changes in mammalian hearts often result in cardiac hypertrophy and fibrosis that are preceded by inflammatory infiltration. In this paper, we show that lifelong treatment of BALB/c and C57BL/6 mice with the mitochondria-targeted antioxidant SkQ1 retards senescence-associated myocardial disease (cardiomyopathy), cardiac hypertrophy, and diffuse myocardial fibrosis. To investigate the molecular basis of the action of SkQ1, we have applied DNA microarray analysis. The global gene expression profile in heart tissues was not significantly affected by administration of SkQ1. However, we found some small but statistically significant modifications of the pathways related to cell-to-cell contact, adhesion, and leukocyte infiltration. Probably, SkQ1-induced decrease in leukocyte and mesenchymal cell adhesion and/or infiltration lead to a reduction in age-related inflammation and subsequent fibrosis. The data indicate a causative role of mitochondrial reactive oxygen species in cardiovascular aging and imply that SkQ1 has potential as a drug against age-related cardiac dysfunction.

The Application of Toxicogenomics for (Drinking) Water Quality Assessment

Due to anthropogenic activities, freshwater systems worldwide are exposed to thousands of compounds. Monitoring of priority pollutants is important, however, monitoring of all individual compounds would be practically impossible. Moreover, the effects on human health remains mostly unknown since compound toxicity data is often absent. With the release of increasing amounts of new (emerging) chemicals into the environment, new monitoring strategies are required to assess the effects of (drinking water relevant) chemicals on human health. Therefore, sensitive in-vitro bioassays have been developed which focus on specific physiological effects such as endocrine disruption or mutagenicity. The advantages of bioassays are that they directly determine the effect of (an entire mixture of) compounds present in an environmental sample instead of identifying single compounds. These bioassays are able to detect known and unknown compounds as long as they trigger the bioassay response. However, a disadvantage of applying these assays may be that they each focus on a (relatively narrow selection of) specific physiological endpoint and that the human relevance is often obscure. The evolution of new technologies and the recent advances in the knowledge on DNA sequences and organisation (i.e. genomics) have enabled the development of new holistic tools, such as DNA microarrays. This report provides an overview of innovative omic methods and the mechanisms behind certain genomics technologies are explained. The main focus is on the potential application of DNA microarrays that are able to measure the impact of toxic substances on gene expression, i.e. transcription of DNA to mRNA (transcriptomics). Practical information on this method, such as sensitivity, analysis time, responsiveness, specificity, etc. is discussed. An overview is presented of the potential applications of this technology with regard to water quality assessment. In addition, the limitations and challenges that need to be overcome are discussed. The following potential applications for genomics in water quality assessment are foreseen: This title belongs to KWR Watercycle Research Institute Series . ISBN: 9781780406671 (eBook)