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  1. Fulltext Mitochondrial DNA integrity and function are critical for endothelium-dependent vasodilation in rats with metabolic syndrome
    Kiyooka T, Ohanyan V, Yin L, Pung YF, Chen YR, Chen CL, et al.
    Basic Res Cardiol, 2022 Jan 17;117(1):3.
    PMID: 35039940 DOI: 10.1007/s00395-021-00908-1
    Endothelial dysfunction in diabetes is generally attributed to oxidative stress, but this view is challenged by observations showing antioxidants do not eliminate diabetic vasculopathy. As an alternative to oxidative stress-induced dysfunction, we interrogated if impaired mitochondrial function in endothelial cells is central to endothelial dysfunction in the metabolic syndrome. We observed reduced coronary arteriolar vasodilation to the endothelium-dependent dilator, acetylcholine (Ach), in Zucker Obese Fatty rats (ZOF, 34 ± 15% [mean ± standard deviation] 10-3 M) compared to Zucker Lean rats (ZLN, 98 ± 11%). This reduction in dilation occurred concomitantly with mitochondrial DNA (mtDNA) strand lesions and reduced mitochondrial complex activities in the endothelium of ZOF versus ZLN. To demonstrate endothelial dysfunction is linked to impaired mitochondrial function, administration of a cell-permeable, mitochondria-directed endonuclease (mt-tat-EndoIII), to repair oxidatively modified DNA in ZOF, restored mitochondrial function and vasodilation to Ach (94 ± 13%). Conversely, administration of a cell-permeable, mitochondria-directed exonuclease (mt-tat-ExoIII) produced mtDNA strand breaks in ZLN, reduced mitochondrial complex activities and vasodilation to Ach in ZLN (42 ± 16%). To demonstrate that mitochondrial function is central to endothelium-dependent vasodilation, we introduced (via electroporation) liver mitochondria (from ZLN) into the endothelium of a mesenteric vessel from ZOF and restored endothelium-dependent dilation to vasoactive intestinal peptide (VIP at 10-5 M, 4 ± 3% vasodilation before mitochondrial transfer and 48 ± 36% after transfer). Finally, to demonstrate mitochondrial function is key to endothelium-dependent dilation, we administered oligomycin (mitochondrial ATP synthase inhibitor) and observed a reduction in endothelium-dependent dilation. We conclude that mitochondrial function is critical for endothelium-dependent vasodilation.
    Matched MeSH terms: DNA, Mitochondrial/metabolism
  2. Hybridisation analysis for serum hepatitis B virus DNA
    Yap SF, Wong PW, Kenneth-Raj
    Br J Biomed Sci, 1994 Dec;51(4):336-40.
    PMID: 7756940
    A study was carried out to determine optimal assay conditions for an in-house hybridisation assay for detection of hepatitis B virus (HBV) genome in serum samples. Pre-treatment of samples, blot treatment and hybridisation conditions were found to affect assay performance. The indirect serum blot procedure was more robust and reliable than direct serum blotting. In the former, viral particles were isolated from the sample, lysed and then extracted. In comparison, no approaches to the direct serum spot method performed adequately. Sensitivity studies showed that labelling of the nucleic acid probe with dCTP was more efficient than with dATP. Using probes labelled to a specific activity of > 1 x 10(8) and an autoradiography period of about 48 h we could achieve a detection limit of < 1 pg. Specificity was achieved by use of a highly purified probe and moderately stringent hybridisation and wash conditions. Background binding was minimal and there was no non-specific binding of probe to negative control samples. Factors affecting speed of the assay were studied to identify steps that could be modified to shorten assay time without sacrificing performance. A shorter centrifugation step and the use of a high specific-activity probe permitted completion of an assay within four days.
    Matched MeSH terms: DNA, Viral/blood*
  3. Fulltext Dispersal direction of Malaysian Fasciola gigantica from neighboring southeast Asian countries inferred using mitochondrial DNA analysis
    Ichikawa-Seki M, Hayashi K, Tashiro M, Khadijah S
    Infect Genet Evol, 2022 Nov;105:105373.
    PMID: 36202207 DOI: 10.1016/j.meegid.2022.105373
    Fasciola gigantica and hybrid Fasciola flukes, responsible for the disease fasciolosis, are found in Southeast Asian countries. In the present study, we performed molecular species identification of Fasciola flukes distributed in Terengganu, Malaysia using multiplex PCR for phosphoenolpyruvate carboxykinase (pepck) and PCR-restriction fragment length polymorphism (RFLP) for DNA polymerase delta (pold). Simultaneously, phylogenetic analysis based on mitochondrial NADH dehydrogenase subunit 1 (nad1) was performed for the first time on Malaysian Fasciola flukes to infer the dispersal direction among neighboring countries. A total of 40 flukes used in this study were identified as F. gigantica. Eight nad1 haplotypes were identified in the F. gigantica population of Terengganu. Median-joining network analysis revealed that the Malaysian population was related to those obtained from bordering countries such as Thailand and Indonesia. However, genetic differentiation was detected using population genetics analyses. Nevertheless, the nucleotide diversity (π) value suggested that F. gigantica with the predominant haplotypes was introduced into Malaysia from Thailand and Indonesia. The dispersal direction suggested by population genetics in the present study may not be fully reliable since Fasciola flukes were collected from a single location in one state of Malaysia. Further studies analyzing more samples from many locations are required to validate the dispersal direction proposed herein.
    Matched MeSH terms: DNA, Mitochondrial/genetics
  4. Fulltext Sequencing crop genomes: a gateway to improve tropical agriculture
    Thottathil, Gincy Paily, Jayasekaran, Kandakumar, Ahmad Sofiman Othman
    Trop Life Sci Res, 2016;27(1):93-114.
    MyJurnal
    Agricultural development in the tropics lags behind development in the
    temperate latitudes due to the lack of advanced technology, and various biotic and abiotic
    factors. To cope with the increasing demand for food and other plant-based products,
    improved crop varieties have to be developed. To breed improved varieties, a better
    understanding of crop genetics is necessary. With the advent of next-generation DNA
    sequencing technologies, many important crop genomes have been sequenced. Primary
    importance has been given to food crops, including cereals, tuber crops, vegetables, and
    fruits. The DNA sequence information is extremely valuable for identifying key genes
    controlling important agronomic traits and for identifying genetic variability among the
    cultivars. However, massive DNA re-sequencing and gene expression studies have to be
    performed to substantially improve our understanding of crop genetics. Application of the
    knowledge obtained from the genomes, transcriptomes, expression studies, and
    epigenetic studies would enable the development of improved varieties and may lead to a
    second green revolution. The applications of next generation DNA sequencing
    technologies in crop improvement, its limitations, future prospects, and the features of
    important crop genome projects are reviewed herein.
    Matched MeSH terms: DNA; Sequence Analysis, DNA
  5. From bites to barcodes: uncovering the hidden diversity of black flies (Diptera: Simuliidae) in Vietnam
    Putt QY, Ya'cob Z, Adler PH, Chen CD, Hew YX, Izwan-Anas N, et al.
    Parasit Vectors, 2023 Aug 07;16(1):266.
    PMID: 37545007 DOI: 10.1186/s13071-023-05892-0
    BACKGROUND: Prompt and precise identification of black flies (Simuliidae) is crucial, given their biting behaviour and significant impact on human and animal health. To address the challenges presented by morphology and chromosomes in black fly taxonomy, along with the limited availability of molecular data pertaining to the black fly fauna in Vietnam, this study employed DNA-based approaches. Specifically, we used mitochondrial and nuclear-encoded genes to distinguish nominal species of black flies in Vietnam.

    METHODS: In this study, 135 mitochondrial cytochrome c oxidase subunit I (COI) sequences were established for 45 species in the genus Simulium in Vietnam, encompassing three subgenera (Gomphostilbia, Nevermannia, and Simulium), with 64 paratypes of 27 species and 16 topotypes of six species. Of these COI sequences, 71, representing 27 species, are reported for the first time.

    RESULTS: Combined with GenBank sequences of specimens from Malaysia, Myanmar, Thailand, and Vietnam, a total of 234 DNA barcodes of 53 nominal species resulted in a 71% success rate for species identification. Species from the non-monophyletic Simulium asakoae, S. feuerborni, S. multistriatum, S. striatum, S. tuberosum, and S. variegatum species groups were associated with ambiguous or incorrect identifications. Pairwise distances, phylogenetics, and species delimitation analyses revealed a high level of cryptic diversity, with discovery of 15 cryptic taxa. The current study also revealed the limited utility of a fast-evolving nuclear gene, big zinc finger (BZF), in discriminating closely related, morphologically similar nominal species of the S. asakoae species group.

    CONCLUSION: This study represents the first comprehensive molecular genetic analysis of the black fly fauna in Vietnam to our knowledge, providing a foundation for future research. DNA barcoding exhibits varying levels of differentiating efficiency across species groups but is valuable in the discovery of cryptic diversity.

    Matched MeSH terms: DNA Barcoding, Taxonomic/methods
  6. Micro-interdigitated electrodes genosensor based on Au-deposited nanoparticles for early detection of cervical cancer
    Jaapar FN, Parmin NA, Halim NHA, Hashim U, Gopinath SCB, Halim FS, et al.
    Int J Biol Macromol, 2023 Dec 31;253(Pt 2):126745.
    PMID: 37689297 DOI: 10.1016/j.ijbiomac.2023.126745
    Genosensor-based electrodes mediated with nanoparticles (NPs) have tremendously developed in medical diagnosis. Herein, we report a facile, rapid, low cost and highly sensitive biosensing strategy for early detection of HPV 18 using gold-nanoparticles (AuNPs) deposited on micro-IDEs. This study represents surface charge transduction of micro-interdigitated electrodes (micro-IDE) alumina insulated with silica, independent and mini genosensor modified with colloidal gold NPs (AuNPs), and determination of gene hybridization for early detection of cervical cancer. The surface of AuNPs deposited micro-IDE functionalized with optimized 3-aminopropyl-triethoxysilane (APTES) followed by hybridization with deoxyribonucleic acid (DNA) virus to develop DNA genosensor. The results of ssDNA hybridization with the ssDNA target of human papillomavirus (HPV) 18 have affirmed that micro-IDE functionalized with colloidal AuNPs resulted in the lowest detection at 0.529 aM. Based on coefficient regression, micro-IDE functionalized with AuNPs produces better results in the sensitivity test (R2 = 0.99793) than unfunctionalized micro-IDE.
    Matched MeSH terms: DNA, Viral/genetics
  7. Specific detection of the Old World screwworm fly, Chrysomya bezziana, in bulk fly trap catches using real-time PCR
    Jarrett S, Morgan JA, Wlodek BM, Brown GW, Urech R, Green PE, et al.
    Med Vet Entomol, 2010 Sep;24(3):227-35.
    PMID: 20497318 DOI: 10.1111/j.1365-2915.2010.00867.x
    The Old World screwworm fly (OWS), Chrysomya bezziana Villeneuve (Diptera: Calliphoridae), is a myiasis-causing blowfly of major concern for both animals and humans. Surveillance traps are used in several countries for early detection of incursions and to monitor control strategies. Examination of surveillance trap catches is time-consuming and is complicated by the presence of morphologically similar flies that are difficult to differentiate from Ch. bezziana, especially when the condition of specimens is poor. A molecular-based method to confirm or refute the presence of Ch. bezziana in trap catches would greatly simplify monitoring programmes. A species-specific real-time polymerase chain reaction (PCR) assay was designed to target the ribosomal DNA internal transcribed spacer 1 (rDNA ITS1) of Ch. bezziana. The assay uses both species-specific primers and an OWS-specific Taqman((R)) MGB probe. Specificity was confirmed against morphologically similar and related Chrysomya and Cochliomyia species. An optimal extraction protocol was developed to process trap catches of up to 1000 flies and the assay is sensitive enough to detect one Ch. bezziana in a sample of 1000 non-target species. Blind testing of 29 trap catches from Australia and Malaysia detected Ch. bezziana with 100% accuracy. The probability of detecting OWS in a trap catch of 50 000 flies when the OWS population prevalence is low (one in 1000 flies) is 63.6% for one extraction. For three extractions (3000 flies), the probability of detection increases to 95.5%. The real-time PCR assay, used in conjunction with morphology, will greatly increase screening capabilities in surveillance areas where OWS prevalence is low.
    Matched MeSH terms: DNA/genetics
  8. Fulltext An Ultrasensitive Voltammetric Genosensor for the Detection of Bacteria Vibrio cholerae in Vegetable and Environmental Water Samples
    Futra D, Tan LL, Lee SY, Lertanantawong B, Heng LY
    Biosensors (Basel), 2023 Jun 04;13(6).
    PMID: 37366981 DOI: 10.3390/bios13060616
    In view of the presence of pathogenic Vibrio cholerae (V. cholerae) bacteria in environmental waters, including drinking water, which may pose a potential health risk to humans, an ultrasensitive electrochemical DNA biosensor for rapid detection of V. cholerae DNA in the environmental sample was developed. Silica nanospheres were functionalized with 3-aminopropyltriethoxysilane (APTS) for effective immobilization of the capture probe, and gold nanoparticles were used for acceleration of electron transfer to the electrode surface. The aminated capture probe was immobilized onto the Si-Au nanocomposite-modified carbon screen printed electrode (Si-Au-SPE) via an imine covalent bond with glutaraldehyde (GA), which served as the bifunctional cross-linking agent. The targeted DNA sequence of V. cholerae was monitored via a sandwich DNA hybridization strategy with a pair of DNA probes, which included the capture probe and reporter probe that flanked the complementary DNA (cDNA), and evaluated by differential pulse voltammetry (DPV) in the presence of an anthraquninone redox label. Under optimum sandwich hybridization conditions, the voltammetric genosensor could detect the targeted V. cholerae gene from 1.0 × 10-17-1.0 × 10-7 M cDNA with a limit of detection (LOD) of 1.25 × 10-18 M (i.e., 1.1513 × 10-13 µg/µL) and long-term stability of the DNA biosensor up to 55 days. The electrochemical DNA biosensor was capable of giving a reproducible DPV signal with a relative standard deviation (RSD) of <5.0% (n = 5). Satisfactory recoveries of V. cholerae cDNA concentration from different bacterial strains, river water, and cabbage samples were obtained between 96.5% and 101.6% with the proposed DNA sandwich biosensing procedure. The V. cholerae DNA concentrations determined by the sandwich-type electrochemical genosensor in the environmental samples were correlated to the number of bacterial colonies obtained from standard microbiological procedures (bacterial colony count reference method).
    Matched MeSH terms: DNA; DNA, Complementary
  9. Fulltext Molecular pathology quality control in Southeast Asia: Results of a multiregional quality assurance study from MASTER KEY Asia
    Okuma HS, Yoshida H, Kobayashi Y, Arakaki M, Mizoguchi C, Inagaki L, et al.
    Cancer Sci, 2023 Jun;114(6):2664-2673.
    PMID: 36919757 DOI: 10.1111/cas.15790
    Tissue specimen quality assurance is a major issue of precision medicine for rare cancers. However, the laboratory standards and quality of pathological specimens prepared in Asian hospitals remain unknown. To understand the methods in Southeast Asian oncology hospitals and to clarify how pre-analytics affect the quality of formalin-fixed paraffin-embedded (FFPE) specimens, a questionnaire surveying pre-analytical procedures (Part I) was administered, quality assessment of immunohistochemistry (IHC) staining and DNA/RNA extracted from the representative FFPE specimens from each hospital (Part II) was conducted, and the quality of DNA/RNA extracted from FFPE of rare-cancer patients for genomic sequencing (Part III) was examined. Quality measurements for DNA/RNA included ΔΔCt, DV200, and cDNA yield. Six major cancer hospitals from Malaysia, Philippines, and Vietnam participated. One hospital showed unacceptable quality for the DNA/RNA assessment, but improved by revising laboratory procedures. Only 57% (n = 73) of the 128 rare-cancer patients' specimens met both DNA and RNA quality criteria for next-generation sequencing. Median DV200 was 80.7% and 64.3% for qualified and failed RNA, respectively. Median ΔΔCt was 1.25 for qualified and 4.89 for failed DNA. Longer storage period was significantly associated with poor DNA (fail to qualify ratio = 1579:321 days, p 
    Matched MeSH terms: DNA/genetics
  10. The use of a molecular technique for the detection of porcine ingredients in the Malaysian food market
    Farouk AE, Batcha MF, Greiner R, Salleh HM, Salleh MR, Sirajudin AR
    Saudi Med J, 2006 Sep;27(9):1397-400.
    PMID: 16951781
    To develop a molecular technique that is fast and reliable in detecting porcine contamination or ingredients in foods.
    Matched MeSH terms: DNA; DNA Fragmentation
  11. Fulltext A novel nano therapeutic using convalescent plasma derived exosomal (CPExo) for COVID-19: A combined hyperactive immune modulation and diagnostics
    Anand K, Vadivalagan C, Joseph JS, Singh SK, Gulati M, Shahbaaz M, et al.
    Chem Biol Interact, 2021 Aug 01;344:109497.
    PMID: 33991505 DOI: 10.1016/j.cbi.2021.109497
    Extracellular vesicles like exosomes are important therapeutic tactics for treating COVID -19. By utilizing convalescent plasma derived exosomes (CPExo) from COVID-19 recovered persistence could accelerate the treatment strategies in the current state of affairs. Adequate literature has shown that administering the exosome to the in vivo system could be beneficial and could target the pathogens in an effective and precise manner. In this hypothesis we highlight the CPExo instead of convalescent plasma (CP), perhaps to dispense of exosomes are gratified and it's more effectively acquired immune response conferral through antibodies. COVID-19 convalescent plasma has billions of exosomes and it has aptitudes to carry molecular constituents like proteins, lipids, RNA and DNA, etc. Moreover, exosomes are capable of recognizing antigens with adequate sensitivity and specificity. Many of these derivatives could trigger an immune modulation into the cells and act as an epigenetic inheritor response to target pathogens through RNAs. COIVID-19 resistance activated plasma-derived exosomes are either responsible for the effects of plasma beyond the contained immune antibodies or could be inhibitory. The proposed hypothesis suggests that preselecting the plasma-derived antibodies and RNAs merged exosomes would be an optimized therapeutic tactic for COVID-19 patients. We suggest that, the CPExo has a multi-potential effect for treatment efficacy by acting as immunotherapeutic, drug carrier, and diagnostic target with noncoding genetic materials as a biomarker.
    Matched MeSH terms: DNA/immunology
  12. Nanoplate-based digital PCR for highly sensitive pork DNA detection targeting multi-copy nuclear and mitochondrial genes
    Mokhtar NFK, Shun YQ, Raja Nhari RMH, Mohamad NA, Shahidan NM, Warsanah IH, et al.
    Food Addit Contam Part A Chem Anal Control Expo Risk Assess, 2024 Feb;41(2):120-133.
    PMID: 38190283 DOI: 10.1080/19440049.2023.2298476
    The inclusion of ingredients derived from pigs in highly processed consumer products poses a significant challenge for DNA-targeted analytical enforcement, which could be overcome by using digital PCR. However, most species detection methods use digital PCR to target single-copy nuclear genes, which limits their sensitivity. In this work, we examined the performance of a nanoplate-based digital PCR method that targets multi-copy nuclear (MPRE42) and mitochondrial (Cytb) genes. Poor separation of positive and negative partitions, as well as a 'rain effect' were obtained in the porcine-specific MPRE42 assay. Among the optimization strategies examined, the inclusion of restriction enzymes slightly improved the separation of positive and negative partitions, but a more extensive 'rain effect' was observed. The high copy number of the MPRE42 amplicon is hypothesized to contribute to the saturation of the positive signal. In contrast, the porcine-specific Cytb assay achieved perfect separation of positive and negative partitions with no 'rain effect'. This assay can detect as little as 0.4 pg of pork DNA, with a sensitivity of 0.05% (w/w) in a pork-chicken mixture, proving its applicability for detecting pork in meat and meat-based products. For the MPRE42 assay, potential applications in highly degraded products such as gelatin and lard are anticipated.
    Matched MeSH terms: DNA/genetics
  13. Preclinical Development of a Novel Epitope-based DNA Vaccine Candidate against SARS-CoV-2 and Evaluation of Immunogenicity in BALB/c Mice
    Khalid K, Lim HX, Anwar A, Tan SH, Hwang JS, Ong SK, et al.
    AAPS PharmSciTech, 2024 Mar 12;25(3):60.
    PMID: 38472523 DOI: 10.1208/s12249-024-02778-x
    The protective efficacies of current licensed vaccines against COVID-19 have significantly reduced as a result of SARS-CoV-2 variants of concern (VOCs) which carried multiple mutations in the Spike (S) protein. Considering that these vaccines were developed based on the S protein of the original SARS-CoV-2 Wuhan strain, we designed a recombinant plasmid DNA vaccine based on highly conserved and immunogenic B and T cell epitopes against SARS-CoV-2 Wuhan strain and the Omicron VOC. Literature mining and bioinformatics were used to identify 6 immunogenic peptides from conserved regions of the SARS-CoV-2 S and membrane (M) proteins. Nucleotide sequences encoding these peptides representing highly conserved B and T cell epitopes were cloned into a pVAX1 vector to form the pVAX1/S2-6EHGFP recombinant DNA plasmid vaccine. The DNA vaccine was intranasally or intramuscularly administered to BALB/c mice and evaluations of humoral and cellular immune responses were performed. The intramuscular administration of pVAX1/S2-6EHGFP was associated with a significantly higher percentage of CD8+ T cells expressing IFN-γ when compared with the empty vector and PBS controls. Intramuscular or intranasal administrations of pVAX1/S2-6EHGFP resulted in robust IgG antibody responses. Sera from mice intramuscularly immunized with pVAX1/S2-6EHGFP were found to elicit neutralizing antibodies capable of SARS-CoV-2 Omicron variant with the ACE2 cell surface receptor. This study demonstrated that the DNA vaccine construct encoding highly conserved immunogenic B and T cell epitopes was capable of eliciting potent humoral and cellular immune responses in mice.
    Matched MeSH terms: Vaccines, DNA*
  14. Detection of somatic mutations in the mitochondrial DNA control region D-loop in brain tumors: The first report in Malaysian patients
    Mohamed Yusoff AA, Mohd Nasir KN, Haris K, Mohd Khair SZN, Abdul Ghani ARI, Idris Z, et al.
    Oncol Lett, 2017 Nov;14(5):5179-5188.
    PMID: 29098023 DOI: 10.3892/ol.2017.6851
    Although the role of nuclear-encoded gene alterations has been well documented in brain tumor development, the involvement of the mitochondrial genome in brain tumorigenesis has not yet been fully elucidated and remains controversial. The present study aimed to identify mutations in the mitochondrial DNA (mtDNA) control region D-loop in patients with brain tumors in Malaysia. A mutation analysis was performed in which DNA was extracted from paired tumor tissue and blood samples obtained from 49 patients with brain tumors. The D-loop region DNA was amplified using the PCR technique, and genetic data from DNA sequencing analyses were compared with the published revised Cambridge sequence to identify somatic mutations. Among the 49 brain tumor tissue samples evaluated, 25 cases (51%) had somatic mutations of the mtDNA D-loop, with a total of 48 mutations. Novel mutations that had not previously been identified in the D-loop region (176 A-deletion, 476 C>A, 566 C>A and 16405 A-deletion) were also classified. No significant associations between the D-loop mutation status and the clinicopathological parameters were observed. To the best of our knowledge, the current study presents the first evidence of alterations in the mtDNA D-loop regions in the brain tumors of Malaysian patients. These results may provide an overview and data regarding the incidence of mitochondrial genome alterations in Malaysian patients with brain tumors. In addition to nuclear genome aberrations, these specific mitochondrial genome alterations may also be considered as potential cancer biomarkers for the diagnosis and staging of brain cancers.
    Matched MeSH terms: DNA Mutational Analysis; DNA, Mitochondrial
  15. Fulltext Phylogeography of the termite Macrotermes gilvus and insight into ancient dispersal corridors in Pleistocene Southeast Asia
    Veera Singham G, Othman AS, Lee CY
    PLoS One, 2017;12(11):e0186690.
    PMID: 29186140 DOI: 10.1371/journal.pone.0186690
    Dispersal of soil-dwelling organisms via the repeatedly exposed Sunda shelf through much of the Pleistocene in Southeast Asia has not been studied extensively, especially for invertebrates. Here we investigated the phylogeography of an endemic termite species, Macrotermes gilvus (Hagen), to elucidate the spatiotemporal dynamics of dispersal routes of terrestrial fauna in Pleistocene Southeast Asia. We sampled 213 termite colonies from 66 localities throughout the region. Independently inherited microsatellites and mtDNA markers were used to infer the phylogeographic framework of M. gilvus. Discrete phylogeographic analysis and molecular dating based on fossil calibration were used to infer the dynamics of M. gilvus dispersal in time and space across Southeast Asia. We found that the termite dispersal events were consistently dated within the Pleistocene time frame. The dispersal pattern was multidirectional, radiating eastwards and southwards out of Indochina, which was identified as the origin for dispersal events. We found no direct dispersal events between Sumatra and Borneo despite the presence of a terrestrial connection between them during the Pleistocene. Instead, central Java served as an important link allowing termite colonies to be established in Borneo and Sumatra. Our findings support the hypothesis of a north-south dispersal corridor in Southeast Asia and suggest the presence of alternative dispersal routes across Sundaland during the Pleistocene. For the first time, we also propose that a west-east dispersal through over-water rafting likely occurred across the Pleistocene South China Sea. We found at least two independent entry routes for terrestrial species to infiltrate Sumatra and Borneo at different times.
    Matched MeSH terms: DNA, Mitochondrial/genetics
  16. Fulltext Building a DNA barcode reference library for the true butterflies (Lepidoptera) of Peninsula Malaysia: what about the subspecies?
    Wilson JJ, Sing KW, Sofian-Azirun M
    PLoS One, 2013;8(11):e79969.
    PMID: 24282514 DOI: 10.1371/journal.pone.0079969
    The objective of this study was to build a DNA barcode reference library for the true butterflies of Peninsula Malaysia and assess the value of attaching subspecies names to DNA barcode records. A new DNA barcode library was constructed with butterflies from the Museum of Zoology, University of Malaya collection. The library was analysed in conjunction with publicly available DNA barcodes from other Asia-Pacific localities to test the ability of the DNA barcodes to discriminate species and subspecies. Analyses confirmed the capacity of the new DNA barcode reference library to distinguish the vast majority of species (92%) and revealed that most subspecies possessed unique DNA barcodes (84%). In some cases conspecific subspecies exhibited genetic distances between their DNA barcodes that are typically seen between species, and these were often taxa that have previously been regarded as full species. Subspecies designations as shorthand for geographically and morphologically differentiated groups provide a useful heuristic for assessing how such groups correlate with clustering patterns of DNA barcodes, especially as the number of DNA barcodes per species in reference libraries increases. Our study demonstrates the value in attaching subspecies names to DNA barcode records as they can reveal a history of taxonomic concepts and expose important units of biodiversity.
    Matched MeSH terms: DNA Barcoding, Taxonomic*
  17. Fulltext Hexaferrocenium tri[hexa(isothiocyanato)iron(III)] trihydroxonium complex as a new DNA intercalator for electrochemical DNA biosensor
    Ariffin EY, Zakariah EI, Ruslin F, Kassim M, Yamin BM, Heng LY, et al.
    Sci Rep, 2021 Apr 12;11(1):7883.
    PMID: 33846405 DOI: 10.1038/s41598-021-86939-z
    Ferrocene or ferrocenium has been widely studied in the field of organometallic complexes because of its stable thermodynamic, kinetic and redox properties. Novel hexaferrocenium tri[hexa(isothiocyanato)iron(III)]trihydroxonium (HexaFc) complex was the product from the reaction of ferrocene, maleic acid and ammonium thiocyanate and was confirmed by elemental analysis CHNS, FTIR and single crystal X-ray crystallography. In this study, HexaFc was used for the first time as an electroactive indicator for porcine DNA biosensor. The UV-Vis DNA titrations with this compound showed hypochromism and redshift at 250 nm with increasing DNA concentrations. The binding constant (Kb) for HexaFc complex towards CT-DNA (calf-thymus DNA) was 3.1 × 104 M-1, indicated intercalator behaviour of the complex. To test the usefulness of this complex for DNA biosensor application, a porcine DNA biosensor was constructed. The recognition probes were covalently immobilised onto silica nanospheres (SiNSs) via glutaraldehyde linker on a screen-printed electrode (SPE). After intercalation with the HexaFc complex, the response of the biosensor to the complementary porcine DNA was measured using differential pulse voltammetry. The DNA biosensor demonstrated a linear response range to the complementary porcine DNA from 1 × 10-6 to 1 × 10-3 µM (R2 = 0.9642) with a limit detection of 4.83 × 10-8 µM and the response was stable up to 23 days of storage at 4 °C with 86% of its initial response. The results indicated that HexaFc complex is a feasible indicator for the DNA hybridisation without the use of a chemical label for the detection of porcine DNA.
    Matched MeSH terms: DNA/analysis*
  18. Fulltext Epigenome-wide association study of adiposity and future risk of obesity-related diseases
    Campanella G, Gunter MJ, Polidoro S, Krogh V, Palli D, Panico S, et al.
    Int J Obes (Lond), 2018 Dec;42(12):2022-2035.
    PMID: 29713043 DOI: 10.1038/s41366-018-0064-7
    BACKGROUND: Obesity is an established risk factor for several common chronic diseases such as breast and colorectal cancer, metabolic and cardiovascular diseases; however, the biological basis for these relationships is not fully understood. To explore the association of obesity with these conditions, we investigated peripheral blood leucocyte (PBL) DNA methylation markers for adiposity and their contribution to risk of incident breast and colorectal cancer and myocardial infarction.

    METHODS: DNA methylation profiles (Illumina Infinium® HumanMethylation450 BeadChip) from 1941 individuals from four population-based European cohorts were analysed in relation to body mass index, waist circumference, waist-hip and waist-height ratio within a meta-analytical framework. In a subset of these individuals, data on genome-wide gene expression level, biomarkers of glucose and lipid metabolism were also available. Validation of methylation markers associated with all adiposity measures was performed in 358 individuals. Finally, we investigated the association of obesity-related methylation marks with breast, colorectal cancer and myocardial infarction within relevant subsets of the discovery population.

    RESULTS: We identified 40 CpG loci with methylation levels associated with at least one adiposity measure. Of these, one CpG locus (cg06500161) in ABCG1 was associated with all four adiposity measures (P = 9.07×10-8 to 3.27×10-18) and lower transcriptional activity of the full-length isoform of ABCG1 (P = 6.00×10-7), higher triglyceride levels (P = 5.37×10-9) and higher triglycerides-to-HDL cholesterol ratio (P = 1.03×10-10). Of the 40 informative and obesity-related CpG loci, two (in IL2RB and FGF18) were significantly associated with colorectal cancer (inversely, P DNA methylation patterns, may be an intermediate biomarker at the intersection of obesity and obesity-related diseases, and could offer clues as to underlying biological mechanisms.

    Matched MeSH terms: DNA Methylation/genetics*
  19. Fulltext DNA, RNA, and protein extraction: the past and the present
    Tan SC, Yiap BC
    J Biomed Biotechnol, 2009;2009:574398.
    PMID: 20011662 DOI: 10.1155/2009/574398
    Extraction of DNA, RNA, and protein is the basic method used in molecular biology. These biomolecules can be isolated from any biological material for subsequent downstream processes, analytical, or preparative purposes. In the past, the process of extraction and purification of nucleic acids used to be complicated, time-consuming, labor-intensive, and limited in terms of overall throughput. Currently, there are many specialized methods that can be used to extract pure biomolecules, such as solution-based and column-based protocols. Manual method has certainly come a long way over time with various commercial offerings which included complete kits containing most of the components needed to isolate nucleic acid, but most of them require repeated centrifugation steps, followed by removal of supernatants depending on the type of specimen and additional mechanical treatment. Automated systems designed for medium-to-large laboratories have grown in demand over recent years. It is an alternative to labor-intensive manual methods. The technology should allow a high throughput of samples; the yield, purity, reproducibility, and scalability of the biomolecules as well as the speed, accuracy, and reliability of the assay should be maximal, while minimizing the risk of cross-contamination.
    Matched MeSH terms: DNA/isolation & purification*
  20. Fulltext Modulation of Cell Cycle Profile by Chlorella vulgaris Prevents Replicative Senescence of Human Diploid Fibroblasts
    Saberbaghi T, Abbasian F, Mohd Yusof YA, Makpol S
    Evid Based Complement Alternat Med, 2013;2013:780504.
    PMID: 23573154 DOI: 10.1155/2013/780504
    In this study, the effects of Chlorella vulgaris (CV) on replicative senescence of human diploid fibroblasts (HDFs) were investigated. Hot water extract of CV was used to treat HDFs at passages 6, 15, and 30 which represent young, presenescence, and senescence ages, respectively. The level of DNA damage was determined by comet assay while apoptosis and cell cycle profile were determined using FACSCalibur flow cytometer. Our results showed direct correlation between increased levels of damaged DNA and apoptosis with senescence in untreated HDFs (P < 0.05). Cell cycle profile showed increased population of untreated senescent cells that enter G0/G1 phase while the cell population in S phase decreased significantly (P < 0.05). Treatment with CV however caused a significant reduction in the level of damaged DNA and apoptosis in all age groups of HDFs (P < 0.05). Cell cycle analysis showed that treatment with CV increased significantly the percentage of senescent HDFs in S phase and G2/M phases but decreased the population of cells in G0/G1 phase (P < 0.05). In conclusion, hot water extract of Chlorella vulgaris effectively decreased the biomarkers of ageing, indicating its potential as an antiageing compound.
    Matched MeSH terms: DNA; DNA Damage
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