Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “molecular identification”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 37 records · Page 2Linked to original sources
PubMed2004

Development of a novel, simple and rapid molecular identification system for clinical Candida species.

Identification of clinical yeast isolates causing candidiasis is routinely performed by commercial yeast identification systems based on biochemical, morphological and physiological tests. These systems require 3-5 days and the proportion of identifications that are incorrect is high. Our novel and rapid molecular identification system for clinical Candida species is based on the analysis of restriction patterns obtained from PCR-generated ribosomal DNA sequences using five restriction enzymes. A software package (CandID) was designed to include a database of restriction fragment length polymorphism (RFLP) patterns for 29 Candida species. For 'in-house' validation, 122 clinical isolates that had previously identified in clinical laboratories were typed by this system. These clinical isolates were also independently re-identified by the API 20C AUX system. The ribosomal DNA RFLP database in the context of supporting analytical software allowed simple and rapid (1 work day) identification.

Candida↗
PubMed1997

Molecular identification of sympatric chromosomal forms of Anopheles gambiae and further evidence of their reproductive isolation.

Three chromosomal forms of Anopheles gambiae s.s., designated as Bamako, Mopti and Savanna, were studied for diagnostic PCR assays based on the analysis of the X-linked ribosomal DNA (rDNA). The study was performed on a 1.3 kb fragment containing part of the 28S coding region and part of the intergenic spacer region. The amplified material was cut with fourteen restriction enzymes to detect Restriction Fragment Length Polymorphisms (RFLPs). The enzymes Tru9I and HhaI produced patterns of DNA bands which differentiated Mopti from Savanna and Bamako; moreover, a distinct 'hybrid' pattern was recognized in the F1 female progeny from the cross of Mopti with either one of the other two chromosomal forms. The diagnostic significance of the PCR-RFLP assay was verified on 203 karyotyped females from field samples collected in two villages in Mali and one village in Burkina Faso. Agreement was observed between the chromosomal and the molecular identifications. No 'hybrid' molecular patterns were detected even among carriers of rare heterokaryotypes hypothetically produced by crosses between Mopti and Savanna. The results confirm previous observations indicating barriers to gene flow within An. gambiae s.s. and supporting the specific status of the taxonomic units proposed on cytogenetic ground.

Animals↗
PubMed2006

Molecular identification and characterization of mannitol-negative methicillin-resistant Staphylococcus aureus.

We report on the isolation, molecular identification, and characterization of 5 mannitol-negative methicillin-resistant Staphylococcus aureus (MRSA) from clinical samples in KwaZulu-Natal (KZN) province, South Africa. Identification based on phenotypic testing and polymerase chain reaction detection of the S. aureus species-specific nuc gene and the coagulase gene indicated that the mannitol-negative isolates were S. aureus. Furthermore, they were mecA positive, and SCCmec typing demonstrated that all the isolates harbored type IV SCCmec. API STAPH (Biomerieux, Marcy-l'Etoile, France) misidentified 2 mannitol-negative MRSA that belonged to the major clone in KZN province, as Staphylococcus lugdunensis. Although the prevalence and mechanism of mannitol-negative MRSA is unknown, laboratories are encouraged to investigate S. aureus with atypical characteristics.

Bacterial Proteins↗
PubMed2005

[Molecular identification of medicinal plants: Dendrobium chrysanthum, Dendrobium fimbriatum and their morphologically allied species by PCR-RFLP analyses].

AIM: To establish a simple method for molecular identification of original plants of D. chrysanthum and D. fimbriatum using molecular marker rDNA ITS region. METHODS: Restriction patterns of ITS fragments were obtained using PCR-RFLP method. The PCR products of D. chrysanthum and its morphologically allied species were digested at 37 degrees C by Cla I and Apa LI, those of D. fimbriatum and its morphologically allied species were digested by Sph I. RESULTS: D. chrysanthum, D. fimbriatum and their morphologically allied species could be identified by predicted restriction profiles of PCR-RFLP. The botanical origin of twenty-five fresh samples of "Shihu" collected in markets was identified by this method. CONCLUSION: The results showed that PCR-RFLP analysis of the rDNA ITS region is a feasible, simple and inexpensive method for determining the botanical origin of the traditional Chinese medicine "Shihu".

DNA, Plant↗
PubMed1986

Electro-magnetic resonance phenomenon as a possible mechanism related to the "bi-digital o-ring test molecular identification and localization method".

The author hypothesizes that the mechanism of the "Bi-Digital O-Ring Test Molecular Identification and Localization Method" is due to an electro-magnetic wave resonance phenomenon between two identical substances having an identical resonance frequency and separated by a known distance. Such a hypothesis was tested and proved by using two identical sets of electro-magnetic resonance circuits, each consisting of a fixed inductance (L) and variable capacitance (C), in place of two identical substances or molecules. When one of the resonance circuits was connected or placed close to the body surface and when the frequency of the other resonance circuit was made identical to the one placed next to the body, a maximal weakening response of the "Bi-Digital O-Ring Test" was observed only when the axes of the coils of the two separate sets of resonance circuits were oriented perpendicular to each other; when the axes of the coils of the two separate sets of resonance circuits were oriented parallel to each other, no "Bi-Digital O-Ring Test" weakening response was observed, even when both resonance circuits had an identical resonance frequency. The information about the molecular structure and quantity of any molecule is contained in the specific electro-magnetic field emitted by the particular molecule. These electro-magnetic waves, containing information about the particular substance, can be propagated through a metal wire, through a "concentrated electro-magnetic field projector," or through a light beam with wavelength longer than green color (particularly a monochromatic, collimated light beam or soft laser beam). In the cases in which a light beam is used, the monochromatic light beam (including laser beam) acts as a very high frequency carrier of the electro-magnetic waves emitted from a particular substance placed near the source of the light beam or near the end of the light beam, and information on the molecular structure and amount of the substance is carried by the light beam (including laser beam) in both forward and backward directions (bi-directional propagation of information). Even reflected light from any molecule or substance in the visual field that reaches the eyes carries information on the substance to the eye ground, particularly when the individual is gazing at the substance; simultaneously, information on a substance not normally existing in the body that happens to be in the body is sent out from the eye ground in electro-magnetic waves to the object being gazed at by the individual.

Electromagnetic Fields↗
PubMed2003

Molecular identification of six medicinal Curcuma plants produced in Sichuan: evidence from plastid trnK gene sequences.

AIM: To establish a rapid and simple molecular identification method for six medicinals: Curcuma: C. longa, C. phaeocaulis, C. sichuanensis, C. chuanyujin, C. chuanhuangjiang, and C. chuanezhu in Sichuan Province. METHODS: A molecular approach (trnK nucleotide sequencing) was used in this study. RESULTS: The sequenced entire chloroplast trnK gene region spanned 2,699-2,705 bp. The matK gene (an intron embodied in trnK gene) sequence and the intron spacer region of the trnK gene have great diversity within these six medicinal Curcuma species. There were six single bases substitutions between trnK coding region and matK region, the 9-bp deletion and 4-bp or 14-bp insertion repeat at some sites of matK region in each taxon. CONCLUSION: These relatively variable sequences were potentially informative in the identification for these six Curcuma species at the DNA level.

Base Sequence↗
PubMed1996

Molecular identification of bacteria from a coculture by denaturing gradient gel electrophoresis of 16S ribosomal DNA fragments as a tool for isolation in pure cultures.

Molecular information about the bacterial composition of a coculture capable of sulfate reduction after exposure to oxic and microoxic conditions was used to identify and subsequently to isolate the components of the mixture in pure culture. PCR amplification of 16S ribosomal DNA fragments from the coculture, analyzed by denaturing gradient gel electrophoresis, resulted in two distinct 16S ribosomal DNA bands, indicating two different bacterial components. Sequencing showed that the bands were derived from a Desulfovibrio strain and an Arcobacter strain. Since the phylogenetic positions of bacteria are often consistent with their physiological properties and culture requirements, molecular identification of the two components of this coculture allowed the design of specific culture conditions to separate and isolate both strains in pure culture. This approach facilitates the combined molecular and physiological analysis of mixed cultures and microbial communities.

Bacteria↗
PubMed2004

Molecular identification of the urea uptake system and transcriptional analysis of urea transporter- and urease-encoding genes in Corynebacterium glutamicum.

The molecular identification of the Corynebacterium glutamicum urea uptake system is described. This ABC-type transporter is encoded by the urtABCDE operon, which is transcribed in response to nitrogen limitation. Expression of the urt genes is regulated by the global nitrogen regulator AmtR, and an amtR deletion strain showed constitutive expression of the urtABCDE genes. The AmtR repressor protein also controls transcription of the urease-encoding ureABCEFGD genes in C. glutamicum. The ure gene cluster forms an operon which is mainly transcribed in response to nitrogen starvation. To confirm the increased synthesis of urease subunits under nitrogen limitation, proteome analyses of cytoplasmic protein extracts from cells grown under nitrogen surplus and nitrogen limitation were carried out, and five of the seven urease subunits were identified.

ATP-Binding Cassette Transporters↗
PubMed2002

[Application of BACTEC MGIT 960 system and molecular identification of mycobacteria in the diagnosis of spinal tuberculosis].

OBJECTIVE: To evaluate the clinical application of BACTEC MGIT 960 system and molecular identification of mycobacterium species in diagnosis of spinal tuberculosis. METHODS: Thirty-one specimens were obtained from patients with spinal tuberculosis, followed by culturing with BACTEC MGIT 960 system and in Löwenstein-Jensen (L-J) medium respectively. Mycobacterium species of the clinical isolates were identified with PCR amplification of IS986 and PCR-single-strand conformation polymorphism (PCR-SSCP) analysis of 16S rRNA, with the conventional identification approach serving as control. RESULTS: The positivity rates of mycobacterium culture with BACTEC MGIT 960 system and L-J medium were 83.87 % and 61.29 % respectively. The average detection time with BACTEC MGIT 960 were 11.3 d, 15.4 d shorter than that with the L-J medium (26.7 d). The genetic identification of mycobacterium species was more rapid and accurate with consistent results with that from conventional method. CONCLUSION: BACTEC MGIT 960 system is preferable for detecting mycobacteria in spinal tuberculosis. The combined application of this instrument with genetic identification of mycobacteria is an effective strategy for the bacteriological diagnosis of tuberculosis.

Adolescent↗
PubMed2000

Molecular identification of cytosolic prostaglandin E2 synthase that is functionally coupled with cyclooxygenase-1 in immediate prostaglandin E2 biosynthesis.

Here we report the molecular identification of cytosolic glutathione (GSH)-dependent prostaglandin (PG) E(2) synthase (cPGES), a terminal enzyme of the cyclooxygenase (COX)-1-mediated PGE(2) biosynthetic pathway. GSH-dependent PGES activity in the cytosol of rat brains, but not of other tissues, increased 3-fold after lipopolysaccharide (LPS) challenge. Peptide microsequencing of purified enzyme revealed that it was identical to p23, which is reportedly the weakly bound component of the steroid hormone receptor/hsp90 complex. Recombinant p23 expressed in Escherichia coli and 293 cells exhibited all the features of PGES activity detected in rat brain cytosol. A tyrosine residue near the N terminus (Tyr(9)), which is known to be critical for the activity of cytosolic GSH S-transferases, was essential for PGES activity. The expression of cPGES/p23 was constitutive and was unaltered by proinflammatory stimuli in various cells and tissues, except that it was increased significantly in rat brain after LPS treatment. cPGES/p23 was functionally linked with COX-1 in marked preference to COX-2 to produce PGE(2) from exogenous and endogenous arachidonic acid, the latter being supplied by cytosolic phospholipase A(2) in the immediate response. Thus, functional coupling between COX-1 and cPGES/p23 may contribute to production of the PGE(2) that plays a role in maintenance of tissue homeostasis.

3T3 Cells↗
PubMed1989

Molecular identification and characterization of the gastrin receptor in guinea pig gastric glands.

The binding characteristics of 125I-labeled Leu15-gastrin and the molecular identification of the gastrin receptor was investigated in dispersed guinea pig gastric glands. The binding of [125I]gastrin to gastric glands was temperature dependent, saturable, and specific. At 37 C, the binding was rapid, became maximal within 10 min, and declined after 30 min; at 24 C, binding reached a steady state between 30 and 60 min. The dissociation of specifically bound gastrin was also rapid, with 35% of the radioligand dissociating in 5 min at 37 C. Gastrin displaced [125I]gastrin in a dose-dependent manner, with 50% displacement at 4.4 nM. Scatchard analysis of the saturation curve was best described by a one-site binding model with a Kd of 2.3 nM and maximum binding of 6.0 x 10(-10) M/mg DNA. A significant reduction of [125I]gastrin binding to glands occurred in the presence of GTP (0.1 mM), (Bu)2-cGMP (1.0 mM), and protease inhibitors. Chemical cross-linking studies using the cross-linking reagent disuccinimidyl suberate and sodium dodecyl sulfate-gel electrophoresis identified a major band with a mol wt of 78K and several other lower mol wt bands in the range of 60K, 48K, and 38K. Identical electrophoretic patterns were obtained when glands were bound and solubilized in the presence and absence of dithiothreitol, showing the lack of disulfide bonds in the gastrin receptor subunit structure. Since dithiothreitol significantly enhanced radioligand binding when present during binding, its observed actions are most likely in the intracellular processing of the radioligand and not at the receptor level.

Animals↗
PubMed2001

cDNA cloning and molecular identification of the major oyster allergen from the Pacific oyster Crassostrea gigas.

BACKGROUND: Shellfish is one of the most common food allergens. Despite the recent cloning and molecular identification of the major heat stable crustacean allergens in shrimp, lobster and crab, there have been no similar studies on molluscs to which a significant portion of populations allergic to shellfish are also hypersensitive. Recent biochemical evidence suggests that tropomyosin is also an allergen in molluscs, but data on the molecular cloning, nucleotide sequencing, expression and IgE binding to mollusc tropomyosin are lacking. OBJECTIVE: This study was undertaken to clone, identify and determine the primary structure of a major IgE-reactive mollusc allergen in oyster at the DNA and protein level. METHODS: We constructed an expression cDNA library from the Pacific oyster Crassostrea gigas. This library was screened for IgE binding clones using sera from 15 subjects with a well-documented history of type I hypersensitivity reactions to oysters. An IgE reactive clone was selected and sub-cloned into plasmids for nucleotide sequence determination and expression in E. coli. RESULTS: We identified a 1.3-kb cDNA designated as Cra g 1.03. Expression of Cra g 1.03 in plasmid vector pGEX produced a 59-kDa recombinant fusion protein reactive to the IgE antibodies from patients with oyster allergies but not non-allergic controls. Cra g 1.03 has an open reading frame of 233 amino acids and demonstrates marked similarity in amino acid composition and peptide sequence with mollusc and crustacean tropomyosins. Absorption of oyster allergic sera with Cra g 1.03 totally removed IgE reactivity to oyster extract. Moreover, absorption of allergic sera with recombinant shrimp tropomyosin (Met e 1), lobster tropomyosin (Pan s 1) and crab tropomyosin (Cha f 1) removed most of the IgE reactivity to Cra g 1.03. CONCLUSION: Cra g 1.03 is the first oyster allergen identified at the molecular level. Nucleotide and amino acid comparison shows that this protein is the oyster tropomyosin.

Allergens↗
PubMed2006

Theory and application of dissociative electron capture in molecular identification.

The coupling of an electron monochromator (EM) to a mass spectrometer (MS) has created a new analytical technique, EM-MS, for the investigation of electrophilic compounds. This method provides a powerful tool for molecular identification of compounds contained in complex matrices, such as environmental samples. In particular, EM-MS has been applied to the detection of nitrated aromatic compounds, many of which are potent mutagens and/or carcinogens and are considered environmental hazards. EM-MS expands the application and selectivity of traditional MS through the inclusion of a new dimension in the space of molecular characteristics-the electron resonance energy spectrum. EM-MS also enhances detection sensitivity as well because the entire electron flux of the proper energy can be delivered into the negative ion resonance that is analytically most useful to solving the problem at hand. However, before this tool can realize its full potential, it will be necessary to create a library of resonance energy scans from standards of the molecules for which EM-MS offers a practical means of detection. Unfortunately, the number of such standards is very large and not all of the compounds are commercially available, making this library difficult to construct. Here, an approach supplementing direct measurement with chemical inference and quantum scattering theory is presented to demonstrate the feasibility of directly calculating resonance energy spectra. This approach makes use of the symmetry of the transition-matrix element of the captured electron to discriminate between the spectra of isomers. As a way of validating this approach, the resonance values for 25 nitrated aromatic compounds were measured along with their relative abundance. Subsequently, the spectra for the isomers of nitrotoluene were shown to be consistent with the symmetry-based model. The initial success of this treatment suggests that it might be possible to predict negative ion resonances and thus create a library of EM-MS standards.

Journal Article↗
PubMed2005

Molecular identification and phylogenetic analysis of baculoviruses from Lepidoptera.

PCR amplification of the highly conserved baculovirus genes late expression factor 8 (lef-8), late expression factor 9 (lef-9) and polyhedrin/granulin (polh/gran) combined with molecular phylogenetic analyses provide a powerful tool to identify lepidopteran-specific baculoviruses and to study their diversity. In the present investigation, we have improved the degenerate oligonucleotides and corroborated the approach that was recently described by Lange et al. (Lange, M., Wang, H., Zhihong, H., Jehle, J.A., 2004. Towards a molecular identification and classification system of lepidopteran-specific baculoviruses. Virology 325, 36-47.). Baculovirus DNA was isolated from 71 uncharacterized historic baculovirus samples, and partial gene sequences were amplified by using gene-specific degenerate PCR primers. The obtained PCR products were directly sequenced, and the deduced amino acid sequences were compiled and aligned with published sequences of these target genes. A phylogenetic tree of 117 baculoviruses was inferred using maximum parsimony and distance methods. Based on the comprehensive phylogenetic analysis of the partial lef-8, lef-9 and polh/gran genes, we propose a phylogenetic species criterion for lepidopteran-specific baculoviruses that uses the genetic distances of these genes for species demarcation.

Animals↗
PubMed2002

Chloride channels and hepatocellular function: prospects for molecular identification.

Hepatocytes possess chloride channels at the plasma membrane and in multiple intracellular compartments. These channels are required for cell volume regulation and acidification of intracellular organelles. Evidence also supports a role of chloride channels in modulation of apoptosis and cell growth. Swelling- and Ca(2+)-activated chloride channels have been identified in hepatocyte plasma membranes, and chloride channels have been observed in the membranes of lysosomes, endosomes, Golgi, endoplasmic reticulum, mitochondria, and the nucleus. This review summarizes the functions of these channels and discusses the specific channel molecules they may represent. Chloride channel molecules shown to be expressed in hepatocytes include members of the ClC channel family (ClC-2, ClC-3, ClC-5, and ClC-7), members of the newly identified CLIC family of intracellular chloride channels (CLIC-1 and CLIC-4), the mitochondrial voltage-dependent anion channel, and a newly identified intracellular channel, MCLC (Mid-1 related chloride channel). Current understanding does not include a molecular identification of most of the observed channel functions, but details of the molecular properties of these channel molecules should allow future identification and further understanding of chloride channel function in hepatocytes.

Animals↗
PubMed2001

Rapid molecular identification and subtyping of Helicobacter pylori by pyrosequencing of the 16S rDNA variable V1 and V3 regions.

We describe here the use of real-time DNA sequence analysis of Helicobacter pylori 16S rRNA gene fragments by pyrosequencing for rapid molecular identification and subtyping of clinical isolates based on DNA sequence heterogeneity within the variable V1 and V3 regions. Six individual 16S rDNA V1 alleles (position 75-100) were identified in 23 clinical isolates obtained from gastric biopsy specimens. Eleven of these revealed sequence identities with H. pylori 26695 and one was identical with the rrn genes in strain J99. The other V1 alleles showing single or double nucleotide mutations or single nucleotide insertions could be divided into four groups with 5, 4, 1, and 1 isolates each. Two out of 25 isolates demonstrated single C to T transitions in the V3 region (position 990-1020). The present findings show that subtle DNA sequence variation occurs sufficiently often in the 16S rDNA variable V1 and V3 regions of H. pylori to provide a consistent system for subtyping.

Bacterial Typing Techniques↗
PubMed1999

Novel application of PhastSystem polyacrylamide gel electrophoresis using restriction fragment length polymorphism--internal transcribed spacer patterns of individuals for molecular identification of entomopathogenic nematodes.

différences! [editorial] [editorial]onomic way of identifying and assigning nematodes to taxons, which had already been determined either by comparative sequence analysis of nuclear rDNA internal transcribed spacer (ITS) region or by other methods of molecular or conventional taxonomy, is provided. Molecular identification of entomopathogenic nematodes (EPN) can be upgraded by basing it on PhastSystem polyacrylamide gel electrophoresis (PAGE) analysis of restriction fragment length polymorphism (RFLP) patterns of polymerase chain reaction (PCR)-amplified DNA derived from single nematodes of Steinernema or Heterorhabditis spp. Although analysis from single worms has previously been made on agarose gel, the resolution on PhastSystem PAGE gel is much higher. The DNA sequences selected for analysis were those constituting the internal transcribed spacer region between the 18S and 26S rDNA genes within the rRNA operon. RFLP analysis was carried out by gel electrophoresis on the PhastSystem (Pharmacia) as detailed elsewhere (Triga et al., Electrophoresis 1999, 20, 1272-1277. The downscaling from conventional agarose to PhastSystem gels resulted in pattern of DNA fragments differing from those obtained with agarose gel electrophoresis under conventional conditions by increasing the number of detected fragments. The approach supported previous species identifications and was able to identify several unclassified isolates, such as those from Hungary and Ireland, and provides a method for identification of previously unclassified strains. We confirmed that Heterorhabditis "Irish Type", represented by two strains of different geographical origin, comprise a species different from H. megidis. We also confirmed that strain IS5 belongs to the species H. indicus rather than to H. bacteriophora, as had been suggested previously.

Animals↗
PubMed2006

Molecular identification of Bacillus thuringiensis var. israelensis to trace its fate after application as a biological insecticide in wetland ecosystems.

AIMS: To determine the fate of viable Bacillus thuringiensis var. israelensis (Bti) spores dispersed in the environment, using a universally applicable molecular detection methodology. METHODS AND RESULTS: Soil samples were spread on growth medium, after a temperature selection of the spores. A PCR amplification of the cry4Aa and cry4Ba insecticidal genes was applied on the colonies. Ribotyping was performed subsequently. This combined molecular method proved to be very specific for Bti, which was easily differentiated from the other B. thuringiensis serovars. A site regularly treated with Vectobac-G was chosen within the 'Bolle di Magadino' natural reserve, and monitored throughout 1 year for the detection of Bti spores. The results showed that the numbers were relatively high after insecticidal applications (1.4 x 10(5) CFU g(-1)), and decreased approx. 10-fold after 220 days. A successive treatment induced a new increase. CONCLUSIONS: The results show that yearly repeated use of Vectobac-G does not seem to have a major ecological impact on the 'Bolle di Magadino' natural reserve. Bti spores followed a trend leading to their eventual disappearance from the ecosystem, despite the seasonal application of this biological insecticide for more than a decade. SIGNIFICANCE AND IMPACT OF THE STUDY: The molecular identification of Bti cells through the PCR analysis of the delta-endotoxins genes coupled to ribotyping, is an innovative method, that has enabled the identification of this organism into wetland environments.

Bacillus thuringiensis↗