Displaying publications 681 - 700 of 3507 in total

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  1. Fulltext A duplex endpoint PCR assay for rapid detection and differentiation of Leptospira strains
    Benacer D, Zain SNM, Lewis JW, Khalid MKNM, Thong KL
    Rev Soc Bras Med Trop, 2017 Mar-Apr;50(2):239-242.
    PMID: 28562762 DOI: 10.1590/0037-8682-0364-2016
    INTRODUCTION:: This study aimed to develop a duplex endpoint PCR assay for rapid detection and differentiation of Leptospira strains.

    METHODS:: Primers were designed to target the rrs (LG1/LG2) and ligB (LP1/LP2) genes to confirm the presence of the Leptospira genus and the pathogenic species, respectively.

    RESULTS:: The assay showed 100% specificity against 17 Leptospira strains with a limit of detection of 23.1pg/µl of leptospiral DNA and sensitivity of 103 leptospires/ml in both spiked urine and water.

    CONCLUSIONS:: Our duplex endpoint PCR assay is suitable for rapid early detection of Leptospira with high sensitivity and specificity.
    Matched MeSH terms: DNA Primers*
  2. Fulltext DNA Methylation of Human Choline Kinase Alpha Promoter-Associated CpG Islands in MCF-7 Cells
    Mohamed Sa'dom SAF, Raikundalia S, Shamsuddin S, See Too WC, Few LL
    Genes (Basel), 2021 06 01;12(6).
    PMID: 34205960 DOI: 10.3390/genes12060853
    Choline kinase (CK) is the enzyme catalyzing the first reaction in CDP-choline pathway for the biosynthesis of phosphatidylcholine. Higher expression of the α isozyme of CK has been implicated in carcinogenesis, and inhibition or downregulation of CKα (CHKA) is a promising anticancer approach. This study aimed to investigate the regulation of CKα expression by DNA methylation of the CpG islands found on the promoter of this gene in MCF-7 cells. Four CpG islands have been predicted in the 2000 bp promoter region of ckα (chka) gene. Six CpG island deletion mutants were constructed using PCR site-directed mutagenesis method and cloned into pGL4.10 vectors for promoter activity assays. Deletion of CpG4C region located between -225 and -56 significantly increased the promoter activity by 4-fold, indicating the presence of important repressive transcription factor binding site. The promoter activity of methylated full-length promoter was significantly lower than the methylated CpG4C deletion mutant by 16-fold. The results show that DNA methylation of CpG4C promotes the binding of the transcription factor that suppresses the promoter activity. Electrophoretic mobility shift assay analysis showed that cytosine methylation at MZF1 binding site in CpG4C increased the binding of putative MZF1 in nuclear extract. In conclusion, the results suggest that DNA methylation decreased the promoter activity by promoting the binding of putative MZF1 transcription factor at CpG4C region of the ckα gene promoter.
    Matched MeSH terms: DNA Methylation*
  3. Fulltext Sex Chromosome Turnover in Bent-Toed Geckos (Cyrtodactylus)
    Keating SE, Blumer M, Grismer LL, Lin A, Nielsen SV, Thura MK, et al.
    Genes (Basel), 2021 01 19;12(1).
    PMID: 33477871 DOI: 10.3390/genes12010116
    Lizards and snakes (squamates) are known for their varied sex determining systems, and gecko lizards are especially diverse, having evolved sex chromosomes independently multiple times. While sex chromosomes frequently turnover among gecko genera, intrageneric turnovers are known only from Gekko and Hemidactylus. Here, we used RADseq to identify sex-specific markers in two species of Burmese bent-toed geckos. We uncovered XX/XY sex chromosomes in Cyrtodactylus chaunghanakwaensis and ZZ/ZW sex chromosomes in Cyrtodactylus pharbaungensis. This is the third instance of intrageneric turnover of sex chromosomes in geckos. Additionally, Cyrtodactylus are closely related to another genus with intrageneric turnover, Hemidactylus. Together, these data suggest that sex chromosome turnover may be common in this clade, setting them apart as exceptionally diverse in a group already known for diverse sex determination systems.
    Matched MeSH terms: Sequence Analysis, DNA/methods
  4. Large BACs transfect more efficiently in circular topology
    Wong YC, Osahor A, Al-Ajli FOM, Narayanan K
    Anal Biochem, 2021 10 01;630:114324.
    PMID: 34363787 DOI: 10.1016/j.ab.2021.114324
    The effect of DNA topology on transfection efficiency of mammalian cells has been widely tested on plasmids smaller than 10 kb, but little is known for larger DNA vectors carrying intact genomic DNA containing introns, exons, and regulatory regions. Here, we demonstrate that circular BACs transfect more efficiently than covalently closed linear BACs. We found up to 3.1- and 8.9- fold higher eGFP expression from circular 11 kb and 100 kb BACs, respectively, compared to linear BACs. These findings provide insights for improved vector development for gene delivery and expression studies of large intact transgenes in mammalian cells.
    Matched MeSH terms: DNA/genetics*
  5. 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*
  6. Development of a T7 RNA polymerase expressing cell line using lentivirus vectors for the recovery of recombinant Newcastle disease virus
    Yeong MY, Cheow PS, Abdullah S, Song AA, Lei-Rossmann J, Tan TK, et al.
    J Virol Methods, 2021 05;291:114099.
    PMID: 33592218 DOI: 10.1016/j.jviromet.2021.114099
    The development of a T7 RNA polymerase (T7 RNAP) expressing cell line i.e. BSR T7/5 cells marks an improvement of reverse genetics for the recovery of recombinant Newcastle disease virus (rNDV). BSR T7/5 is developed by transient transfection of plasmid encoding T7 RNAP gene for rNDV rescue. However, the gene expression decreases gradually over multiple passages and eventually hinders the rescue of rNDV. To address this issue, lentiviral vector was used to develop T7 RNAP-expressing HEK293-TA (HEK293-TA-Lv-T7) and SW620 (SW620-Lv-T7) cell lines, evidenced by the expression of T7 RNAP after subsequent 20 passages. rNDV was rescued successfully using HEK293-TA-Lv-T7 clones (R1D3, R1D8, R5B9) and SW620-Lv-T7 clones (R1C11, R3C5) by reverse transfection, yielding comparable virus rescue efficiency and virus titres to that of BSR T7/5. This study provides new tools for rNDV rescue and insights into cell line development and virology by reverse genetics.
    Matched MeSH terms: DNA-Directed RNA Polymerases/genetics
  7. Flea-borne pathogens in the cat flea Ctenocephalides felis and their association with mtDNA diversity of the flea host
    Azrizal-Wahid N, Sofian-Azirun M, Low VL
    Comp Immunol Microbiol Infect Dis, 2021 Apr;75:101621.
    PMID: 33609991 DOI: 10.1016/j.cimid.2021.101621
    Flea-borne pathogens were screened from 100 individual cat fleas using a PCR approach, of which 38 % were infected with at least one bacterium. Overall, 28 % of the flea samples were positive for Bartonella as inferred from ITS DNA region. Of these, 25 % (7/28) were identified as Bartonella clarridgeiae, 42.9 % (12/28) as Bartonella henselae consisted of two different strains, and 32.1 % (9/28) as Bartonella koehlerae, which was detected for the first time in Malaysia. Sequencing of gltA amplicons detected Rickettsia DNA in 14 % of cat flea samples, all of them identified as Rickettsia asembonensis (100 %). None of the flea samples were positive for Mycoplasma DNA in 16S rRNA gene detection. Four fleas were co-infected with Bartonella and Rickettsia DNAs. Statistical analyses reveal no significant association between bacterial infection and mtDNA diversity of the cat flea. Nevertheless, in all types of pathogen infections, infected populations demonstrated lower nucleotide and haplotype diversities compared to uninfected populations. Moreover, lower haplotype numbers were observed in infected populations.
    Matched MeSH terms: DNA, Mitochondrial/genetics*
  8. The nearly complete mitogenome of the Southeast Asian firefly Pteroptyx tener (Coleoptera: Lampyridae)
    Cheng S, Mat-Isa MN, Sapian IS, Ishak SF
    Mol Biol Rep, 2021 Feb;48(2):1281-1290.
    PMID: 33582950 DOI: 10.1007/s11033-021-06189-0
    The estuarine firefly, Pteroptyx tener, aggregates in the thousands in mangrove trees lining tidal rivers in Southeast Asia where they engage one another in a nocturnal, pre-mating ritual of synchronised courtship flashes. Unfortunately, populations of the species by virtue of being restricted to isolated estuarine rivers systems in the region, are at risk of genetic isolation. Because of this concern we undertook the task of sequencing and characterising the mitochondrial DNA genome of P. tener, as the first step towards helping us to characterise and better understand their genetic diversity. We sequenced and assembled the mitochondrial DNA genome of P. tener from two male and female specimens from the district of Kuala Selangor in Peninsular Malaysia and announce the molecules in this publication. We also reconstructed the phylogenetic trees of all available lampyrids mitogenomes and suggest the need to re-examine our current understanding of their classification which have largely been based on morphological data and the cox1 gene. Separately, our analysis of codon usage patterns among lampyrid mitogenomes showed that the codon usage in a majority of the protein-coding genes were non-neutral. Codon usage patterns between mitogenome sequences of P. tener were, however, largely neutral. Our findings demonstrate the usefulness of mitochondrial genes/mitogenomes for analysing both inter- and intra- specific variation in the Lampyridae to aid in species discovery in this highly variable genus; and elucidate the phylogenetic relationships of Pteroptyx spp. from the region.
    Matched MeSH terms: DNA, Mitochondrial/genetics*
  9. Integration of advanced technologies for plant variety and cultivar identification
    Azizi MMF, Lau HY, Abu-Bakar N
    J Biosci, 2021;46.
    PMID: 34544910
    Identification of plant variety and cultivar is pivotal in the agricultural sector due to the abundance of plant varieties and cultivars developed in many crop species. However, plant variety and cultivar identification via basic morphological features is problematic and challenging when differentiating closely related species not only due to their limited differences but also due to technical limitations of the process being time-consuming, labour-intensive and costly, and statistically imprecise information being available due to phenotypic plasticity. Therefore, it is imperative to have rapid and highly efficient techniques to mitigate these limitations. This review provides an overview and summarization of the development and application of molecular markers such as Random Amplified Polymorphic DNA (RAPD), Restriction Fragment Length Polymorphism (RFLP), Simple Sequence Repeats (SSR), Inter-simple sequence repeats (ISSR), Amplified Fragment Length Polymorphism (AFLP), Single nucleotide polymorphism (SNP) and DNA barcoding, High-resolution melting (HRM) and biosensor technology as potential tools in the identification of plant variety and cultivar.
    Matched MeSH terms: DNA, Plant/genetics*
  10. Fulltext Species-specific identification of porcine blood plasma in heat-treated chicken meatballs
    Shahimi S, Abd Mutalib S, Ismail N, Elias A, Hashim H, Kashim MIAM
    Saudi J Biol Sci, 2021 Apr;28(4):2447-2452.
    PMID: 33911957 DOI: 10.1016/j.sjbs.2021.01.043
    This study was conducted to detect the presence of chicken and porcine DNA in meatballs using mitochondria DNA (mtDNA) of cytochrome b (cyt b) and nuclear DNA (nDNA) short interspersed nuclear element (SINE) species-specific primers, respectively. While, the mtDNA primers targeted transfer RNA-ATP8 (tRNA-ATP8) gene was used for 1 and 5% (w/w) chicken meatball spiked with commercial porcine blood plasm. Chicken meatballs spiked with 1% and 5% (v/w) fresh and commercial porcine blood plasma, respectively were prepared and heat-treated using five (n = 5) cooking methods: boiling, pan-frying, roasting, microwaving and autoclaving. Two pairs of mtDNA and nDNA primers used, produced 129 and 161 bp amplicons, respectively. Whereas, tRNA-ATP8 primers produced 212 bp of amplicon. Electrophoresis analysis showed positive results for porcine DNA at 1% and 5% (w/w or v/v) for all of the different cooking techniques, either for fresh or commercial blood plasma using SINE primers but not for tRNA-ATP8 primers. The present study has highlighted the useful of species-specific primers of SINE primers in PCR analysis for detecting porcine DNA blood plasma in heat-treated chicken meatballs.
    Matched MeSH terms: DNA, Mitochondrial; DNA Primers
  11. Fulltext Integrated Approach for Species Identification and Quality Analysis for Labisia pumila Using DNA Barcoding and HPLC
    Tarmizi AAA, Wagiran A, Mohd Salleh F, Chua LS, Abdullah FI, Hasham R, et al.
    Plants (Basel), 2021 Apr 07;10(4).
    PMID: 33917172 DOI: 10.3390/plants10040717
    Labisia pumila is a precious herb in Southeast Asia that is traditionally used as a health supplement and has been extensively commercialized due to its claimed therapeutic properties in boosting a healthy female reproductive system. Indigenous people used these plants by boiling the leaves; however, in recent years it has been marketed as powdered or capsuled products. Accordingly, accuracy in determination of the authenticity of these modern herbal products has faced great challenges. Lack of authenticity is a public health risk because incorrectly used herbal species can cause adverse effects. Hence, any measures that may aid product authentication would be beneficial. Given the widespread use of Labisia herbal products, the current study focuses on authenticity testing via an integral approach of DNA barcoding and qualitative analysis using HPLC. This study successfully generated DNA reference barcodes (ITS2 and rbcL) for L. pumila var. alata and pumila. The DNA barcode that was generated was then used to identify species of Labisia pumila in herbal medicinal products, while HPLC was utilized to determine their quality. The findings through the synergistic approach (DNA barcode and HPLC) implemented in this study indicate the importance of both methods in providing the strong evidence required for the identification of true species and to examine the authenticity of such herbal medicinal products.
    Matched MeSH terms: DNA; DNA Barcoding, Taxonomic
  12. 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
  13. Fulltext Sequencing and Annotated of Goat's Milk using Bioinformatics Tool: ClustalX
    Muhammad Najmi Mohammad Fauzi, Aisyah Aqilah Abu Bakar, Liyana Amalina Adnan, Tg Ainul Farha Tg Abdul Rahman, A’wani Aziz Nurdalila
    Malaysian Journal of Science, Health & Technology, 2021;7(1):28-33.
    MyJurnal
    Bioinformatics tool is a software program made to extract meaningful information from the mass of molecular biology or biological databases and carry out sequence or structural analysis. The method of determining the order of nucleotides within a deoxyribonucleic acid (DNA) molecule is known as DNA sequencing. This analysis is meant to be run to the commercialized or factorymade goat's milk (pasteurised) from various states in Malaysia to identify the milk's authenticity, either it is pure or mixed with other foreign substances from other animals. The main objective is to compare DNA sequences of commercialized and raw goat's milk (handmilking and non-pasteurised). To achieve this, we used ClustalX to align and compare the obtained DNA from both milk samples. The sequences will be aligned using ClustalX software. ClustalX is a provider of an automated system for performing multiple alignments of sequences and profiles and evaluating the outcomes. The usage of ClustalX is helpful as it is cost-effective, user-friendly, and showing a high accuracy of the analysis.

    Matched MeSH terms: DNA; Sequence Analysis, DNA
  14. Evaluation of the Roche MagNA Pure 96 nucleic acid extraction platform for the Seegene Anyplex II HPV28 detection assay
    Atchison S, Shilling H, Balgovind P, Machalek DA, Hawkes D, Garland SM, et al.
    J Appl Microbiol, 2021 Nov;131(5):2592-2599.
    PMID: 33942451 DOI: 10.1111/jam.15126
    AIM: Validate the Roche, MagNAPure96 (MP96) nucleic acid extraction platform for Seegene Anyplex II HPV28 (Anyplex28) detection of Human Papillomavirus.

    METHODS AND RESULTS: Comparisons were made for Anyplex28 genotyping from 115 cervical samples extracted on the Hamilton, STARlet and the MP96. Two DNA concentrations were used for the MP96, one matched for sample input to the STARlet and another 5× concentration (laboratory standard). Agreement of HPV detection was 89·8% (κ = 0·798; P = 0·007), with HPV detected in 10 more samples for the MP96. There was a high concordance of detection for any oncogenic HPV genotype (κ = 0·77; P = 0·007) and for any low-risk HPV genotype (κ = 0·85; P = 0·008). DNA extracted at laboratory standard had a lower overall agreement 85·2% (κ = 0·708; P DNA with a higher analytical sensitivity on the Anyplex28.

    SIGNIFICANCE AND IMPACT OF THE STUDY: This analysis supports the use of samples extracted on the MP96 for HPV genotyping using the Anyplex28. Furthermore, an increase in DNA concentration increased analytical sensitivity of the Anyplex28, particularly appropriate for prevalence studies.

    Matched MeSH terms: DNA, Viral/genetics
  15. Fulltext CYTOCHROME OXIDASE I (COI) DIVERGENCE ASSESSMENT OF FAMILY NEMIPTERIDAE FROM MALAYSIAN WATERS
    LIEW YOU EN, SALWANI ABDULLAH, TAN MIN PAU, MAZLAN ABD GHAFFAR, ALIAS MAN, TUN NURUL AIMI MAT JAAFAR
    UMT Journal of Undergraduate Research, 2019;1(1):41-48.
    MyJurnal
    DNA Barcoding, primarily focusing on cytochrome coxidase subunit I (COI) gene has been appraised as an effective tool for species identification. Nonetheless, species identification based on molecular approach is essential for discrimination of look-alike species. In this study, we focused on the marine fishes Family Nemipteridae, one of the commercially important group distributed within the surrounding seas of Malaysia. Some of the samples were collected during the National Demersal Trawl Survey in the Exclusive Economic Zone of East Coast Peninsular Malaysia by the Department of Fishery Malaysia. A 652bp region of COI was sequenced for 74 individuals from nine putative species. Additional 34 COIsequences from GenBank were also included in this study making the total number of samples analysed to 108 individuals. The averageKimura 2-parameter (K2P) nucleotide divergence was 0.34% among individuals within species and 6.97% within genera. All putative species formed monophyletic clades in both Neighbour-joining (NJ) and Maximum-likelihood (ML) trees. However, there was a potential misidentification in specimen identified as Nemipterus tambuloides,as the specimen did not group with their own taxa. It was genetically grouped in Nemipterus thosaporni clade. This study supports the effectiveness of COIgene in species discrimination of Family Nemipteridae.
    Matched MeSH terms: DNA; DNA Barcoding, Taxonomic
  16. Sulfitobacter alexandrii sp. nov., a new microalgae growth-promoting bacterium with exopolysaccharides bioflocculanting potential isolated from marine phycosphere
    Yang Q, Ge YM, Iqbal NM, Yang X, Zhang XL
    Antonie Van Leeuwenhoek, 2021 Jul;114(7):1091-1106.
    PMID: 33895907 DOI: 10.1007/s10482-021-01580-0
    Marine phycosphere harbors unique cross-kingdom associations with enormous ecological significance in aquatic ecosystems as well as relevance for algal biotechnology industry. During our investigating the microbial composition and bioactivity of marine phycosphere microbiota (PM), a novel lightly yellowish and versatile bacterium designated strain AM1-D1T was isolated from cultivable PM of marine dinoflagellate Alexandrium minutum amtk4 that produces high levels of paralytic shellfish poisoning toxins (PSTs). Strain AM1-D1T demonstrates notable bioflocculanting bioactivity with bacterial exopolysaccharides (EPS), and microalgae growth-promoting (MGP) potential toward its algal host. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain AM1-D1T was affiliated to the members of genus Sulfitobacter within the family Rhodobacteraceae, showing the highest sequence similarity of 97.9% with Sulfitobacter noctilucae NB-68T, and below 97.8% with other type strains. The complete genome of strain AM1-D1T consisted of a circular 3.84-Mb chromosome and five circular plasmids (185, 95, 15, 205 and 348 Kb, respectively) with the G+C content of 64.6%. Low values obtained by phylogenomic calculations on the average nucleotide identity (ANI, 77.2%), average amino acid identity (AAI, 74.7%) and digital DNA-DNA hybridization (dDDH, 18.6%) unequivocally separated strain AM1-D1T from its closest relative. The main polar lipids were identified as phosphatidylglycerol, phosphatidylethanolamine, phosphatidylcholine, diphosphatidylglycerol, one unidentified phospholipid and one unidentified lipid. The predominant fatty acids (> 10%) were C18:1 ω7c, C19:0 cyclo ω8c and C16:0. The respiratory quinone was Q-10. The genome of strain AM1-D1T was predicted to encode series of gene clusters responsible for sulfur oxidation (sox) and utilization of dissolved organic sulfur exometabolites from marine dinoflagellates, taurine (tau) and dimethylsulfoniopropionate (DMSP) (dmd), as well as supplementary vitamin B12 (cob), photosynthesis carotenoids (crt) which are pivotal components during algae-bacteria interactions. Based on the evidences by the polyphasic characterizations, strain AM1-D1T represents a novel species of the genus Sulfitobacter, for which the name Sulfitobacter alexandrii sp. nov. is proposed. The type strain is AM1-D1T (= CCTCC 2017277T = KCTC 62491T).
    Matched MeSH terms: DNA, Bacterial; Sequence Analysis, DNA
  17. 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
  18. Isolation and characterization of microsatellite markers and analysis of genetic variability in Curculigo latifolia Dryand
    Babaei N, Abdullah NA, Saleh G, Abdullah TL
    Mol Biol Rep, 2012 Nov;39(11):9869-77.
    PMID: 22752726
    Curculin, a sweet protein found in Curculigo latifolia fruit has great potential for the pharmaceutical industry. This protein interestingly has been found to have both sweet taste and taste-modifying capacities comparable with other natural sweeteners. According to our knowledge this is the first reported case on the isolation of microsatellite loci in this genus. Hence, the current development of microsatellite markers for C. latifolia will facilitate future population genetic studies and breeding programs for this valuable plant. In this study 11 microsatellite markers were developed using 3' and 5' ISSR markers. The primers were tested on 27 accessions from all states of Peninsular Malaysia. The number of alleles per locus ranged from three to seven, with allele size ranging from 141 to 306 bp. The observed and expected heterozygosity ranged between 0.00-0.65 and 0.38-0.79, respectively. The polymorphic information content ranged from 0.35 to 0.74 and the Shannon's information index ranged from 0.82 to 1.57. These developed polymorphic microsatellites were used for constructing a dendrogram by unweighted pair group method with arithmetic mean cluster analysis using the Dice's similarity coefficient. Accessions association according to their geographical origin was observed. Based on characteristics of isolated microsatellites for C. latifolia accessions all genotype can be distinguished using these 11 microsatellite markers. These polymorphic markers could also be applied to studies on uniformity determination and somaclonal variation of tissue culture plantlets, varieties identification, genetic diversity, analysis of phylogenetic relationship, genetic linkage maps and quantitative trait loci in C. latifolia.
    Matched MeSH terms: DNA, Plant/chemistry
  19. Polymorphic microsatellite loci from an indigenous Asian fungus-growing termite, Macrotermes gilvus (Blattodea: Termitidae) and cross amplification in related taxa
    Singham GV, Vargo EL, Booth W, Othman AS, Lee CY
    Environ Entomol, 2012 Apr;41(2):426-31.
    PMID: 22507019 DOI: 10.1603/EN11228
    The fungus-growing termite, Macrotermes gilvus (Hagen), an indigenous species from Southeast Asia distributed from Myanmar to Indonesia and the Philippines, offers great potential as an ecological model system to elucidate the effects of geography on gene flow within this region. We used next generation sequencing (Roche 454 pyrosequencing) to identify microsatellite markers from the genomic DNA of M. gilvus. A modest sequencing volume generated 34,122 reads, with 1,212 (3.6%) reads contains microsatellites with di-, tri-, tetra-, penta-, and hexa-nucleotide repeat motifs. Thirty-seven loci were selected for primer development and tested for polymorphism across 22 colonies of M. gilvus. Eleven loci were found to be polymorphic with 2-4 alleles per locus. Observed and expected heterozygosities ranged between 0.091-0.727 and 0.090-0.540, respectively. Cross taxa amplification was successful across a panel of four related termite species and four multiplex groups were designed for future population genetic studies. These markers will open new avenues for the study of phylogeography and population genetics of this fungus-growing termite. This study also has effectively demonstrated the use of 454 pyrosequencing for the rapid development of informative microsatellite markers from a termite genome.
    Matched MeSH terms: Sequence Analysis, DNA/methods
  20. Isolation by the 5' anchored PCR technique and characterization of eighteen microsatellite loci in horseshoe crab (Tachypleus gigas)
    Ling LP, Adibah AB, Tan SG, Christianus A, Faridah QZ
    J Genet, 2011 Dec;90(3):e101-4.
    PMID: 22232191
    Matched MeSH terms: DNA Primers/chemistry
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