12 research outputs found
Sequence specific recognition of ssDNA by fluorophore 3-hydroxyflavone (vol 153, pg 391, 2015)
No full textSequence specific recognition of ssDNA by fluorophore 3-hydroxyflavone
No full textA fully water soluble 3-hydroxyflavone (3HF) derivative, N-(3-hydroxy-4'-flavonyl)-N,N,N-trimethylammonium sulfate (3HFNMe3) was synthesized. Investigation of its emissions at varying wavelengths revealed that it had three emission bands of normal (N*), anionic (A*) and tautomeric (T*), in ultrapure water. Recognition of single-stranded ten ssDNA chains, having different nucleotide sequences was studied, using the ratiometric change of the intensities of the two bands (A*/T*), depending upon the varying environment of the 3HFNMe3 with different ssDNA chains. Addition of the ssDNA chains to the 3HFNMe3 solution caused gradual quenching of the A* band and had almost no effect on the T*. band. As the ratios of the two bands (A*/T*) vs increasing amount of the ssDNAs generated characteristic curves for each ssDNA chain, it became possible to identify the chains with their characteristic curves. (C) 2015 Elsevier B.V. All rights reserved
Comparison of DNA extraction methods for meat analysis
No full textPreventing adulteration of meat and meat products with less desirable or objectionable meat species is important not only for economical, religious and health reasons, but also, it is important for fair trade practices, therefore, several methods for identification of meat and meat products have been developed. In the present study, ten different DNA extraction methods, including Tris-EDTA Method, a modified Cetyltrimethylammonium Bromide (CTAB) Method, Alkaline Method, Urea Method, Salt Method, Guanidinium Isothiocyanate (GuSCN) Method, Wizard Method, Qiagen Method, Zymogen Method and Genespin Method were examined to determine their relative effectiveness for extracting DNA from meat samples. The results show that the salt method is easy to perform, inexpensive and environmentally friendly. Additionally, it has the highest yield among all the isolation methods tested. We suggest this method as an alternative method for DNA isolation from meat and meat products. (C) 2016 Elsevier Ltd. All rights reserved
Hidroksisinamik Asit Türevlerinin Canlı-Dışı Helicobacter Pylori Karşıtı Etkileri ile Üreaz Enzimini Engelleme Etkinliklerinin Araştırılması
No full textÖZET
Helicobacter pylori (H. pylori), insanlarda, gastrit, peptik ülser, gastrik kanser ve mukoza ilintili lenfoid doku lenfoması gibi ciddi mide
hastalıklarına yol açan önemli bir hastalık etkenidir. H. pylori, ürettiği üreaz enzimleri sayesinde üreyi, karbondioksit ve amonyağa parçalayarak
mide çeperinin asidik koşullarını normal pH’ya getirip hayatta kalabildiği için bu bakterilerle vücudun savaşımı kolay değildir. H.
pylori enfeksiyonları için antibiyotikler mevcut olsa da antibiyotik direnci gelişimi nedeniyle bu tedaviler sonuçsuz kalabilmekte ve yeni
antibiyotiklere gereksinim her geçen gün artmaktadır.
Hidroksisinamik asit türevleri basit fenolik asitler olup meyvelerde, meyve çekirdeklerinde ve sebzelerde bulunmaktadır. Ferulik asit, kafeik
asit, p-kumarik asit, sinapik asit, sözü edilen bu fenolik asit grubuna ait olup antioksidan, anti-inflamatuvar, antimikrobiyal özelliklere
sahiptir ve bu nedenle bazı bakteri enfeksiyonlarının tedavisinde, ilaçlara seçenek olarak kullanılmaktadır. Bu çalışmamızda, bu üç fenolik
asidin H. pylori üzerindeki antimikrobiyal etkinliği ve ürez enzimini engelleme etkisi araştırıldı.
Hidroksisinamik asit türevlerinin anti-H. pylori etkisi H. pylori G27 standart suşu üzerinde test edildi. Minimum inhibisyon konsantrasyonu
(MİK), değerleri 512 ila 0,5 ug/mL arasında değiştiği seri tüp seyreltme yöntemiyle, minimum bakterisidal konsantrasyon (MBK) değerleri
ise MİK içinde kullanılan aynı konsantrasyonlarda canlı ve ölü bakterilerin nispi oranının hesaplanması ile belirlendi. MİK için CLSI M07-A9,
MBC için CLSI M26-A protokolleri kullanıldı. Ureaz inhibisyon aktivitesi Helicheck, üreaz aktivitesine özgü indikatörlü besiyerinde ölçülmüştür.
H. pylori’ye karşı test edilen tüm bileşikler için MİK 64 ug/mL ve MBK 128 ug/mL idi. Test edilen bileşiklerin H. pylori tarafından salgılanan üreaz
enzimi üzerinde hiçbir inhibisyonu saptanmadı. Nükleotid salma deneyi sonuçlarına göre, hidroksisinamik asit türevlerinin bakteri zarında
hasara sebep olması ile zarda oluşan deliklerden dışarı salınma eğiliminde olması beklenen herhangi bir nüklotit miktarı ölçülememiştir.
Gerçekleştirilen bu çalışma, literatür taramalarımız doğrulutusunda, hidroksisinamik asit türevlerinin anti-H. pylori aktivitesini gösteren
ilk çalışmadır. Bu bileşiklerin anti-H. pylori üzerindeki etki mekanizmasını anlamak için daha ileri analizlere ihtiyaç vardır
A simple guanidinium isothiocyanate method for bacterial genomic DNA isolation
No full textA high-quality and low-cost genomic DNA isolation method is needed for use in microbiology laboratories. In this study, we developed a new modified guanidinium isothiocyanate method to isolate DNA from Escherichia coli, Salmonella enterica subsp. enterica serotype Typhimurium, and Staphylococcus aureus and compared it with 4 other DNA isolation methods. The results show that the modified guanidinium isothiocyanate method developed in our laboratory is simple, fast, and inexpensive and yields DNA whose quality and quantity are similar to that of 2 commercial extraction kits but far better than 2 conventional DNA extraction methods used for comparison.1125-1293
Comparison of DNA extraction methods for meat analysis
No full textPreventing adulteration of meat and meat products with less desirable or objectionable meat species is important not only for economical, religious and health reasons, but also, it is important for fair trade practices, therefore, several methods for identification of meat and meat products have been developed. In the present study, ten different DNA extraction methods, including Tris-EDTA Method, a modified Cetyltrimethylammonium Bromide (CTAB) Method, Alkaline Method, Urea Method, Salt Method, Guanidinium Isothiocyanate (GuSCN) Method, Wizard Method, Qiagen Method, Zymogen Method and Genespin Method were examined to determine their relative effectiveness for extracting DNA from meat samples. The results show that the salt method is easy to perform, inexpensive and environmentally friendly. Additionally, it has the highest yield among all the isolation methods tested. We suggest this method as an alternative method for DNA isolation from meat and meat products. (C) 2016 Elsevier Ltd. All rights reserved
Syntheses and evaluation of multicaulin and miltirone-like compounds as antituberculosis agents
No full textFour multicaulin and miltirone-like phenanthrene derivatives were synthesised and evaluated as antituberculosis agents. The crucial step of the synthesis was Pschorr coupling of 4-(3-isopropyl-4-methoxyphenyl)-2-(2-aminophenyl)ethane (13) to give 2-isopropyl-3-methoxy-9,10-dihydrophenanthrene (9) and 4-isopropyl-3-methoxy-9,10-dihydrophenanthrene (9a). Compound 9 was converted to multicaulin and miltirone-like phenanthrene derivatives by further reactions. The best antituberculosis activity was exhibited by 2-isopropylphenanthrene-3-ol (11)
Effective Immobilization of Novel Antimicrobial Peptides via Conjugation onto Activated Silicon Catheter Surfaces
No full textAntibiotic-resistant microorganisms have become a serious threat to public health, resulting in hospital infections, the majority of which are caused by commonly used urinary tract catheters. Strategies for preventing bacterial adhesion to the catheters’ surfaces have been potentially shown as effective methods, such as coating thesurface with antimicrobial biomolecules. Here, novel antimicrobial peptides (AMPs) were designed as potential biomolecules to prevent antibiotic-resistant bacteria from binding to catheter surfaces. Thiolated AMPs were synthesized using solid-phase peptide synthesis (SPPS), and prep-HPLC was used to obtain AMPs with purity greater than 90%. On the other side, the silicone catheter surface was activated by UV/ozone treatment, followed by functionalization with allyl moieties for conjugation to the free thiol group of cystein in AMPs using thiol-ene click chemistry. Peptide-immobilized surfaces were found to become more resistant to bacterial adhesion while remaining biocompatible with mammalian cells. The presence and site of conjugation of peptide molecules were investigated by immobilizing them to catheter surfaces from both ends (C-Pep and Pep-C). It was clearly demonstrated that AMPs conjugated to the surface via theirN terminus have a higher antimicrobial activity. This strategy stands out for its effective conjugation of AMPs to silicone-based implant surfaces for the elimination of bacterial infections
Inter-laboratory assessment of different digital PCR platforms for quantification of human cytomegalovirus DNA
No full textQuantitative PCR (qPCR) is an important tool in pathogen detection. However, the use of different qPCR components, calibration materials and DNA extraction methods reduces comparability between laboratories, which can result in false diagnosis and discrepancies in patient care. The wider establishment of a metrological framework for nucleic acid tests could improve the degree of standardisation of pathogen detection and the quantification methods applied in the clinical context. To achieve this, accurate methods need to be developed and implemented as reference measurement procedures, and to facilitate characterisation of suitable certified reference materials. Digital PCR (dPCR) has already been used for pathogen quantification by analysing nucleic acids. Although dPCR has the potential to provide robust and accurate quantification of nucleic acids, further assessment of its actual performance characteristics is needed before it can be implemented in a metrological framework, and to allow adequate estimation of measurement uncertainties. Here, four laboratories demonstrated reproducibility (expanded measurement uncertainties below 15%) of dPCR for quantification of DNA from human cytomegalovirus, with no calibration to a common reference material. Using whole-virus material and extracted DNA, an intermediate precision (coefficients of variation below 25%) between three consecutive experiments was noted. Furthermore, discrepancies in estimated mean DNA copy number concentrations between laboratories were less than twofold, with DNA extraction as the main source of variability. These data demonstrate that dPCR offers a repeatable and reproducible method for quantification of viral DNA, and due to its satisfactory performance should be considered as candidate for reference methods for implementation in a metrological framework
International Comparison of Enumeration-Based Quantification of DNA Copy-Concentration Using Flow Cytometric Counting and Digital Polymerase Chain Reaction
No full textEnumeration-based determination of DNA copy-concentration was assessed through an international comparison among national metrology institutes (NMIs) and designated institutes (DIs). Enumeration-based quantification does not require a calibration standard thereby providing a route to "absolute quantification", which offers the potential for reliable value assignments of DNA reference materials, and International System of Units (SI) traceability to copy number 1 through accurate counting. In this study, 2 enumeration based methods, flow cytometric (FCM) counting and the digital polymerase chain reaction (dPCR), were compared to quantify a solution of the pBR322 plasmid at a concentration of several thousand copies per microliter. In addition, 2 orthogonal chemical-analysis methods based on nucleotide quantification, isotope-dilution mass spectrometry (IDMS) and capillary electrophoresis (CE) were applied to quantify a more concentrated solution of the plasmid. Although 9 dPCR results from 8 laboratories showed some dispersion (relative standard deviation [RSD] = 11.8%), their means were closely aligned with those of the FCM-based counting method and the orthogonal chemical-analysis methods, corrected for gravimetric dilution factors. Using the means of dPCR results, the RSD of all 4 methods was 1.8%, which strongly supported the validity of the recent enumeration approaches. Despite a good overall agreement, the individual dPCR results were not sufficiently covered by the reported measurement uncertainties. These findings suggest that some laboratories may not have considered all factors contributing to the measurement uncertainty of dPCR, and further investigation of this possibility is warranted
