A rapid and sensitive approach to mutation detection using real-time polymerase chain reaction and melting curve analyses, using BRCA1 as an example.
AuthorsPals G,Pindolia K,Worsham MJ
JournalMolecular diagnosis : a journal devoted to the understanding of human disease through the clinical application of molecular biology
PubMed ID10553024
Comparison of multiple DNA dyes for real-time PCR: effects of dye concentration and sequence composition on DNA amplification and melting temperature.
AuthorsGudnason H,Dufva M,Bang DD,Wolff A
JournalNucleic acids research
PubMed ID17897966
The importance of real-time polymerase chain reaction (PCR) has increased steadily in clinical applications over the last decade. Many applications utilize SYBR Green I dye to follow the accumulation of amplicons in real time. SYBR Green I has, however, a number of limitations that include the inhibition of PCR, preferential ... More
Diagnosis of herpes simplex virus infections in the clinical laboratory by LightCycler PCR.
Herpes simplex virus (HSV) causes several clinical manifestations in both normal and immunocompromised hosts; this agent is the most frequently detected virus in diagnostic laboratories. Recovery of the virus in cell culture is considered the âgold standardâ for detection of this virus from sources other than cerebrospinal fluid. LightCycler is ... More
Kruppel-like factor 2 (KLF2) regulates proinflammatory activation of monocytes.
JournalProceedings of the National Academy of Sciences of the United States of America
PubMed ID16617118
The mechanisms regulating activation of monocytes remain incompletely understood. Herein we provide evidence that Kruppel-like factor 2 (KLF2) inhibits proinflammatory activation of monocytes. In vitro, KLF2 expression in monocytes is reduced by cytokine activation or differentiation. Consistent with this observation, KLF2 expression in circulating monocytes is reduced in patients with ... More
Characterization of a new NIH-registered variant human embryonic stem cell line, BG01V: a tool for human embryonic stem cell research.
Influence of DNA target melting behavior on real-time PCR quantification.
AuthorsWilhelm J,Hahn M,Pingoud A
JournalClinical chemistry
PubMed ID11067807
DNA melting analysis for detection of single nucleotide polymorphisms.
AuthorsLipsky RH,Mazzanti CM,Rudolph JG,Xu K,Vyas G,Bozak D,Radel MQ,Goldman D
JournalClinical chemistry
PubMed ID11274012
Antisense inhibition of hyaluronan synthase-2 in human articular chondrocytes inhibits proteoglycan retention and matrix assembly.
AuthorsNishida Y,Knudson CB,Nietfeld JJ,Margulis A,Knudson W
JournalThe Journal of biological chemistry
PubMed ID10419509
Detection of duck hepatitis B virus DNA on filter paper by PCR and SYBR green dye-based quantitative PCR.
AuthorsWang CY,Giambrone JJ,Smith BF
JournalJournal of clinical microbiology
PubMed ID12089280
Duck hepatitis B virus (DHBV) belongs to the Hepadnaviridae family, which includes human Hepatitis B virus (HBV) and Woodchuck hepatitis virus. It is widely distributed in wild and domestic ducks due to congenital transmission. HBV is a worldwide health problem, with carriers at risk of developing cirrhosis and liver cancer. ... More
Cloning of the multipartite promoter of the sodium-calcium exchanger gene NCX1 and characterization of its activity in vascular smooth muscle cells.
AuthorsScheller T, Kraev A, Skinner S, Carafoli E
JournalJ Biol Chem
PubMed ID9516469
The sodium-calcium exchange activity is mediated by proteins encoded in a small gene family, of which the gene NCX1 is ubiquitously expressed in mammalian tissues. In this study, the multipartite promoter of this gene was analyzed in the human and rat genomes by means of DNA cloning, reverse transcriptase-polymerase chain ... More
Alteration of pharmacokinetics of cyclophosphamide and suppression of the cytochrome p450 genes by ciprofloxacin.
AuthorsXie HJ, Broberg U, Griskevicius L, Lundgren S, Carlens S, Meurling L, Paul C, Rane A, Hassan M
JournalBone Marrow Transplant
PubMed ID12621481
Recently, it has been reported that prophylactic administration of ciprofloxacin during cyclophosphamide (CY) conditioning was a high-risk factor for relapse in patients undergoing allogeneic BMT. In the present study, we investigated the possible mechanisms of this interaction in male Sprague-Dawley rats. The kinetics of CY and its active 4-OH-CY metabolite ... More
cAMP-Dependent protein kinase (PKA) subunit mRNA levels in postmortem brain from patients with bipolar affective disorder (BD).
AuthorsChang A, Li PP, Warsh JJ
JournalBrain Res Mol Brain Res
PubMed ID12941458
Earlier findings of elevated basal and stimulated PKA activities, and increased immunoreactive levels of PKA regulatory and catalytic subunits in discrete postmortem brain regions from bipolar disorder (BD) patients suggest that disturbances in PKA are involved in the pathophysiology of BD. PKA subunit mRNA levels were measured using SYBR Green ... More
Human DNA quantitation using Alu element-based polymerase chain reaction.
Human forensic casework requires sensitive quantitation of human nuclear DNA from complex sources. Widely used commercially available systems detect both nonhuman and human primate DNA, often require special equipment, and have a detection limit of approximately 0.1ng. Multicopy Alu elements include recently integrated subfamilies that are present in the human ... More
Quantitative intra-short interspersed element PCR for species-specific DNA identification.
AuthorsWalker JA, Hughes DA, Anders BA, Shewale J, Sinha SK, Batzer MA
JournalAnal Biochem
PubMed ID12711348
We have designed and evaluated four assays based upon PCR amplification of short interspersed elements (SINEs) for species-specific detection and quantitation of bovine, porcine, chicken, and ruminant DNA. The need for these types of approaches has increased drastically in response to the bovine spongiform encephalopathy epidemic. Using SYBR Green-based detection, ... More
Real-time PCR methods for independent quantitation of TTV and TLMV.
AuthorsMoen EM, Sleboda J, Grinde B
JournalJ Virol Methods
PubMed ID12020793
There is considerable interest in the possible clinical effects of the human circoviruses TT virus (TTV) and TTV-like mini virus (TLMV). Most people appear to have at least one of these viruses replicating actively in their bodies, thus mere correlation of the presence of virus and disease states are probably ... More
A novel strategy for human papillomavirus detection and genotyping with SybrGreen and molecular beacon polymerase chain reaction.
AuthorsSzuhai K, Sandhaus E, Kolkman-Uljee SM, Lemaître M, Truffert JC, Dirks RW, Tanke HJ, Fleuren GJ, Schuuring E, Raap AK
JournalAm J Pathol
PubMed ID11696426
Human papillomaviruses (HPVs) play an important role in the pathogenesis of cervical cancer. For identification of the large number of different HPV types found in (pre)malignant lesions, a robust methodology is needed that combines general HPV detection with HPV genotyping. We have developed for formaldehyde-fixed samples a strategy that, in ... More
Neuronal RNA granules: a link between RNA localization and stimulation-dependent translation.
AuthorsKrichevsky AM, Kosik KS
JournalNeuron
PubMed ID11719208
RNA granules are a macromolecular structure observed in neurons, where they serve as motile units that translocate mRNAs. Isolated RNA granules are highly enriched in Staufen protein and ultrastructurally contain densely packed clusters of ribosomes. With depolarization, many mRNAs, including those involved in plasticity, rapidly shift from the RNA granule ... More
Quantitative detection of Neospora caninum in bovine aborted fetuses and experimentally infected mice by real-time PCR.
AuthorsCollantes-Fernández E, Zaballos A, Alvarez-GarcÃa G, Ortega-Mora LM
JournalJ Clin Microbiol
PubMed ID11923330
We report the development of a real-time PCR assay for the quantitative detection of Neospora caninum in infected host tissues. The assay uses the double-stranded DNA-binding dye SYBR Green I to continuously monitor product formation. Oligonucleotide primers were designed to amplify a 76-bp DNA fragment corresponding to the Nc5 sequence ... More
Demonstration of preferential binding of SYBR Green I to specific DNA fragments in real-time multiplex PCR.
AuthorsGiglio S, Monis PT, Saint CP
JournalNucleic Acids Res
PubMed ID14602929
SYBR Green I (SG) is widely used in real-time PCR applications as an intercalating dye and is included in many commercially available kits at undisclosed concentrations. Binding of SG to double-stranded DNA is non-specific and additional testing, such as DNA melting curve analysis, is required to confirm the generation of ... More
Comparison of real-time PCR with SYBR Green I or 5'-nuclease assays and dot-blot hybridization with rDNA-targeted oligonucleotide probes in quantification of selected faecal bacteria.
AuthorsMalinen E, Kassinen A, Rinttilä T, Palva A
JournalMicrobiology
PubMed ID12576600
PCR primers and hybridization probes were designed for the 16S rRNA genes of six bacterial species or groups typically present in human faeces or used in the dairy industry. The primers and probes were applied for quantification of the target bacterial genomes added in artificial DNA mixtures or faecal DNA ... More
Real-time quantitative polymerase chain reaction diagnosis of infectious posterior uveitis.
AuthorsDworkin LL, Gibler TM, Van Gelder RN
JournalArch Ophthalmol
PubMed ID12427068
OBJECTIVE: To validate a real-time polymerase chain reaction (PCR) assay allowing rapid and sensitive detection and quantitation of 4 common infectious posterior uveitis pathogens. METHODS: A real-time PCR assay using previously validated primer sets for cytomegalovirus, herpes simplex virus, varicella-zoster virus, and Toxoplasma gondii was developed. A standard curve for ... More
Real-time PCR for rapid genotyping of angiotensin-converting enzyme insertion/deletion polymorphism.
OBJECTIVE: To develop a real-time PCR technique for detection of the insertion/deletion (I/D) polymorphism of angiotensin-converting enzyme (ACE) gene. DESIGN AND METHODS: Three primers were designed for performing real-time PCR in the presence of SYBR Green I as flurochrome followed by melting curve analysis. Forty human genomic DNA that have ... More
In-tube DNA methylation profiling by fluorescence melting curve analysis.
AuthorsWorm J, Aggerholm A, Guldberg P
JournalClin Chem
PubMed ID11427447
BACKGROUND: Most PCR assays for detection of 5-methylcytosine in genomic DNA entail a two-step procedure, comprising initial PCR amplification and subsequent product analysis in separate operations that usually require manual transfer. These methods generally provide information about methylation of only a few CpG dinucleotides within the target sequence. METHODS: An ... More
Optical thermal cycler for use as a fluorimetric plate reader to estimate DNA concentrations.
AuthorsGonzalez JM, Saiz-Jimenez C
JournalBiotechniques
PubMed ID12703293
Hairpin primers for simplified single-nucleotide polymorphism analysis of Mycobacterium tuberculosis and other organisms.
AuthorsHazbón MH, Alland D
JournalJ Clin Microbiol
PubMed ID15004082
We describe a novel, simple, rapid, and highly sensitive method to detect single-nucleotide polymorphisms (SNPs) in Mycobacterium tuberculosis and other organisms. Amplification refractory mutation (ARMS) SNP assays were modified by converting the SNP-detecting linear primers in the ARMS assay to hairpin-shaped primers (HPs) through the addition of a 5' tail ... More
Rapid detection of Vibrio vulnificus in shellfish and Gulf of Mexico water by real-time PCR.
AuthorsPanicker G, Myers ML, Bej AK
JournalAppl Environ Microbiol
PubMed ID14711681
In this paper we describe optimization of SYBR Green I-based real-time PCR parameters and testing of a large number of microbial species with vvh-specific oligonucleotide primers to establish a rapid, specific, and sensitive method for detection of Vibrio vulnificus in oyster tissue homogenate and Gulf of Mexico water (gulf water). ... More
Development of melting temperature-based SYBR Green I polymerase chain reaction methods for multiplex genetically modified organism detection.
AuthorsHernández M, RodrÃguez-Lázaro D, Esteve T, Prat S, Pla M
JournalAnal Biochem
PubMed ID14656521
Commercialization of several genetically modified crops has been approved worldwide to date. Uniplex polymerase chain reaction (PCR)-based methods to identify these different insertion events have been developed, but their use in the analysis of all commercially available genetically modified organisms (GMOs) is becoming progressively insufficient. These methods require a large ... More
Simultaneous detection of enteric viruses by multiplex real-time RT-PCR.
AuthorsBeuret C
JournalJ Virol Methods
PubMed ID14656455
A multiplex real-time RT-PCR protocol for the simultaneous detection of noroviruses ("Norwalk-like viruses") of genogroups I and II, human astroviruses and enteroviruses is described. The protocol was developed and evaluated using the LightCycler and corresponding SYBR Green reagents. New primers were designed within conserved genome regions to optimize the detection ... More
Rapid analysis of the DNA-binding specificities of transcription factors with DNA microarrays.
AuthorsMukherjee S, Berger MF, Jona G, Wang XS, Muzzey D, Snyder M, Young RA, Bulyk ML
JournalNat Genet
PubMed ID15543148
'We developed a new DNA microarray-based technology, called protein binding microarrays (PBMs), that allows rapid, high-throughput characterization of the in vitro DNA binding-site sequence specificities of transcription factors in a single day. Using PBMs, we identified the DNA binding-site sequence specificities of the yeast transcription factors Abf1, Rap1 and Mig1. ... More
Involvement of endonuclease G in nucleosomal DNA fragmentation under sustained endogenous oxidative stress.
AuthorsIshihara Y, Shimamoto N
JournalJ Biol Chem
PubMed ID16407272
'We have previously shown that inhibition of catalase and glutathione peroxidase activities by 3-amino-1,2,4-triazole (ATZ) and mercaptosuccinic acid (MS), respectively, in rat primary hepatocytes caused sustained endogenous oxidative stress and apoptotic cell death without caspase-3 activation. In this study, we investigated the mechanism of this apoptotic cell death in terms ... More
Role of diacylglycerol kinase alpha in the attenuation of receptor signaling.
AuthorsSanjuán MA, Jones DR, Izquierdo M, Mérida I
JournalJ Cell Biol
PubMed ID11285286
'Diacylglycerol kinase (DGK) is suggested to attenuate diacylglycerol-induced cell responses through the phosphorylation of this second messenger to phosphatidic acid. Here, we show that DGKalpha, an isoform highly expressed in T lymphocytes, translocates from cytosol to the plasma membrane in response to two different receptors known to elicit T cell ... More
Conserved role of a complement-like protein in phagocytosis revealed by dsRNA knockout in cultured cells of the mosquito, Anopheles gambiae.
AuthorsLevashina EA, Moita LF, Blandin S, Vriend G, Lagueux M, Kafatos FC
JournalCell
PubMed ID11257225
'We characterize a novel hemocyte-specific acute phase glycoprotein from the malaria vector, Anopheles gambiae. It shows substantial structural and functional similarities, including the highly conserved thioester motif, to both a central component of mammalian complement system, factor C3, and to a pan-protease inhibitor, alpha2-macroglobulin. Most importantly, this protein serves as ... More
Molecular cloning of mouse type 2 and type 3 inositol 1,4,5-trisphosphate receptors and identification of a novel type 2 receptor splice variant.
AuthorsIwai M, Tateishi Y, Hattori M, Mizutani A, Nakamura T, Futatsugi A, Inoue T, Furuichi T, Michikawa T, Mikoshiba K
JournalJ Biol Chem
PubMed ID15632133
'We isolated cDNAs encoding type 2 and type 3 inositol 1,4,5-trisphosphate (IP(3)) receptors (IP(3)R2 and IP(3)R3, respectively) from mouse lung and found a novel alternative splicing segment, SI(m2), at 176-208 of IP(3)R2. The long form (IP(3)R2 SI(m2)(+)) was dominant, but the short form (IP(3)R2 SI(m2)(-)) was detected in all tissues ... More
High-resolution detection of loss of heterozygosity of dinucleotide microsatellite markers.
AuthorsHourihan RN, O'Sullivan GC, Morgan JG
JournalBiotechniques
PubMed ID11233603
'Dinucleotide microsatellite markers are frequently investigated to study inheritance, genetic stability, and allele frequency distribution in a wide variety of genetic disorders. Previous studies have encountered significant problems regarding resolution and detection of dinucleotide, microsatellites. In this study, a useful method to investigate loss of heterozygosity (LOH) of dinucleotide microsatellite ... More
Application of differential inflammatory cell count as a tool to monitor udder health.
AuthorsPillai SR, Kunze E, Sordillo LM, Jayarao BM
JournalJ Dairy Sci
PubMed ID11417700
'A flow cytometric technique called differential inflammatory cell count was standardized by staining bovine peripheral blood leukocytes with a combination of DNA binding dyes SYBR green 1 and propidium iodide in water. Leukocytes were also stained with propidium iodide in detergent to determine total cell count. Differential inflammatory cell count ... More
'The ability to multiplex PCR by probe color and melting temperature (T(m)) greatly expands the power of real-time analysis. Simple hybridization probes with only a single fluorescent dye can be used for quantification and allele typing. Different probes are labeled with dyes that have unique emission spectra. Spectral data are ... More
Quantitative analysis of gene amplification in insecticide-resistant Culex mosquitoes.
AuthorsPaton MG, Karunaratne SH, Giakoumaki E, Roberts N, Hemingway J
JournalBiochem J
PubMed ID10657234
'The amplification of carboxylesterase structural genes followed by their overexpression is the most common mechanism of resistance to organophosphorus insecticides in Culex mosquitoes. Most resistant Culex quinquefasciatus mosquitoes have co-amplified estalpha2(1) and estbeta2(1) genes. Recently, Southern, DNA dot-blot analysis and phosphorimaging technology were used to quantify the est gene copy ... More
Leptin induces angiopoietin-2 expression in adipose tissues.
AuthorsCohen B, Barkan D, Levy Y, Goldberg I, Fridman E, Kopolovic J, Rubinstein M
JournalJ Biol Chem
PubMed ID11152449
'Adipose tissues consisting of adipocytes, microvasculature, and stroma are completely ablated upon over-expression of leptin in rats. This tissue regression is mediated by enhanced lipid beta-oxidation, adipocyte dedifferentiation, and apoptosis. To further characterize this phenomenon, we studied the possible effect of leptin on the adipose microvasculature. Tissue microvasculature is maintained ... More
DOTAP cationic liposomes prefer relaxed over supercoiled plasmids.
AuthorsEven-Chen S, Barenholz Y
JournalBiochim Biophys Acta
PubMed ID11118529
'Cationic liposomes and DNA interact electrostatically to form complexes called lipoplexes. The amounts of unbound (free) DNA in a mixture of cationic liposomes and DNA at different cationic lipid:DNA molar ratios can be used to describe DNA binding isotherms; these provide a measure of the binding efficiency of DNA to ... More
Overexpression of WISP-1 down-regulated motility and invasion of lung cancer cells through inhibition of Rac activation.
AuthorsSoon LL, Yie TA, Shvarts A, Levine AJ, Su F, Tchou-Wong KM
JournalJ Biol Chem
PubMed ID12529380
'Wnt-induced-secreted-protein-1 (WISP-1) is a cysteine-rich, secreted factor belonging to the CCN family. These proteins have been implicated in the inhibition of metastasis; however, the mechanisms involved have not been described. We demonstrated that overexpression of WISP-1 in H460 lung cancer cells inhibited lung metastasis and in vitro cell invasion and ... More
Multiplex LightCycler PCR assay for detection and differentiation of Bordetella pertussis and Bordetella parapertussis in nasopharyngeal specimens.
AuthorsSloan LM, Hopkins MK, Mitchell PS, Vetter EA, Rosenblatt JE, Harmsen WS, Cockerill FR, Patel R
JournalJ Clin Microbiol
PubMed ID11773099
'A rapid real-time multiplex PCR assay for detecting and differentiating Bordetella pertussis and Bordetella parapertussis in nasopharyngeal swabs was developed. This assay (LC-PCR-IS) targets the insertion sequences IS481 and IS1001 of B. pertussis and B. parapertussis, respectively, and is performed using the LightCycler (Roche Molecular Biochemicals, Indianapolis, Ind.). The analytical ... More
Ultrasensitive detection of genetically modified maize DNA by capillary gel electrophoresis with laser-induced fluorescence using different fluorescent intercalating dyes.
AuthorsGarcía-Cañas V, González R, Cifuentes A
JournalJ Agric Food Chem
PubMed ID12137467
'In this work, four different fluorescent intercalating dyes are compared for the ultrasensitive CGE-LIF detection of DNA from transgenic maize in flours. The fluorescent intercalating dyes compared are YOPRO-1, SYBR-Green-I, Ethidium bromide (EthBr), and EnhanCE. For all the four dyes optimum concentrations are established, and efficient separations of DNA fragments ... More
Optimization of procedures for counting viruses by flow cytometry.
AuthorsBrussaard CP
JournalAppl Environ Microbiol
PubMed ID15006772
'The development of sensitive nucleic acid stains, in combination with flow cytometric techniques, has allowed the identification and enumeration of viruses in aquatic systems. However, the methods used in flow cytometric analyses of viruses have not been consistent to date. A detailed evaluation of a broad range of sample preparations ... More
Real-time polymerase chain reaction for Mycoplasma gallisepticum in chicken trachea.
AuthorsCarli KT, Eyigor A
JournalAvian Dis
PubMed ID14562901
'In this work, we describe a rapid detection procedure for Mycoplasma gallisepticum from chicken tracheal swabs by real-time polymerase chain reaction (PCR) by LightCycler system, where we were able to monitor the amplification of the newly synthesized M. gallisepticum-specific PCR product as a proportionally increasing fluorescent signal by using the ... More
Real-time PCR-based genotyping assay for CXCR2 polymorphisms.
AuthorsGupta M, Song P, Yates CR, Meibohm B
JournalClin Chim Acta
PubMed ID14967163
'BACKGROUND: The human chemokine receptor CXCR2 (IL8RB) is a high affinity receptor for interleukin-8 as well as other CXC chemokines, and is involved in the chemotaxis of immune cells. Genetic variants of CXCR2 have potential relevance in various inflammatory human disorders. We developed a real-time polymerase chain reaction (PCR)-based allelic ... More
Identification of genes differentially expressed by nerve growth factor- and neurotrophin-3-dependent sensory neurons.
AuthorsFriedel RH, Schnürch H, Stubbusch J, Barde YA
JournalProc Natl Acad Sci U S A
PubMed ID9356508
'The neurotrophins nerve growth factor (NGF) and neurotrophin-3 (NT3) support the survival of subpopulations of primary sensory neurons with defined and distinct physiological characteristics. Only a few genes have been identified as being differentially expressed in these subpopulations, and not much is known about the nature of the molecules involved ... More
Identification of Leptospira biflexa by real-time homogeneous detection of rapid cycle PCR product.
AuthorsWoo TH, Patel BK, Cinco M, Smythe LD, Norris MA, Symonds ML, Dohnt MF, Piispanen J
JournalJ Microbiol Methods
PubMed ID10076627
'Sequence analysis of 16S rRNA genes extracted from nucleic acids databases enabled the identification of a Leptospira biflexa (L. biflexa) signature sequence, against which a reverse primer designated L613, was designed. This primer, when used in conjunction with a universal bacterial specific forward primer designated Fd1, enabled the development of ... More
Quantification of mRNA using real-time RT-PCR.
AuthorsNolan T, Hands RE, Bustin SA
JournalNat Protoc
PubMed ID17406449
'The real-time reverse transcription polymerase chain reaction (RT-qPCR) addresses the evident requirement for quantitative data analysis in molecular medicine, biotechnology, microbiology and diagnostics and has become the method of choice for the quantification of mRNA. Although it is often described as a "gold" standard, it is far from being a ... More
Growth rate regulation of rRNA content of a marine synechococcus (Cyanobacterium) strain
AuthorsBinder BJ, Liu YC
JournalAppl Environ Microbiol
PubMed ID9726881
'The relationship between growth rate and rRNA content in a marine Synechococcus strain was examined. A combination of flow cytometry and whole-cell hybridization with fluorescently labeled 16S rRNA-targeted oligonucleotide probes was used to measure the rRNA content of Synechococcus strain WH8101 cells grown at a range of light-limited growth rates. ... More
A quantitative fluorescence-based microplate assay for the determination of double-stranded DNA using SYBR Green I and a standard ultraviolet transilluminator gel imaging system.
'Various assays are available for quantification of DNA in solution, but none has been described that is both sensitive and specific for double-stranded (ds) DNA and features practical properties such as low dye and equipment costs, speed, and highly parallel microplate formats. Here we show that quantitative and sensitive measurement ... More
Application of digital image analysis and flow cytometry to enumerate marine viruses stained with SYBR gold.
AuthorsChen F, Lu JR, Binder BJ, Liu YC, Hodson RE
JournalAppl Environ Microbiol
PubMed ID11157214
'A novel nucleic acid stain, SYBR Gold, was used to stain marine viral particles in various types of samples. Viral particles stained with SYBR Gold yielded bright and stable fluorescent signals that could be detected by a cooled charge-coupled device camera or by flow cytometry. The fluorescent signal strength of ... More
A high-throughput method for detection of DNA in chloroplasts using flow cytometry.
AuthorsRowan BA, Oldenburg DJ, Bendich AJ
JournalPlant Methods
PubMed ID17381841
'BACKGROUND: The amount of DNA in the chloroplasts of some plant species has been shown recently to decline dramatically during leaf development. A high-throughput method of DNA detection in chloroplasts is now needed in order to facilitate the further investigation of this process using large numbers of tissue samples. RESULTS: ... More
Monitoring phytoplankton, bacterioplankton, and virioplankton in a coastal inlet (Bedford Basin) by flow cytometry.
AuthorsLi WK, Dickie PM
JournalCytometry
PubMed ID11429774
'BACKGROUND: To establish the prevailing state of the ecosystem for the assessment of long-term change, the abundance of microbial plankton in Bedford Basin (Nova Scotia, Canada) is monitored weekly by flow cytometry. METHODS: Phytoplankton are detected by their chlorophyll autofluorescence. Those that contain phycoerythrin are designated as Synechococcus cyanobacteria or ... More
Potential of LightCycler technology for quantification of minimal residual disease in childhood acute lymphoblastic leukemia.
AuthorsEckert C, Landt O, Taube T, Seeger K, Beyermann B, Proba J, Henze G
JournalLeukemia
PubMed ID10673751
'A certain quantity of residual leukemic cells at several time points during chemotherapy of childhood acute lymphoblastic leukemia (ALL) was proved to predict outcome. Future childhood ALL treatment will take minimal residual disease (MRD) into consideration for stratification aiming at an individualization of chemotherapeutic regimens. Recently, the first quantitative real-time ... More
Evaluation of a p30 gene-based real-time reverse transcriptase polymerase chain reaction assay for detection of feline caliciviruses.
'This report describes a feline calicivirus (FCV) p30 gene-based real-time SYBR Green I reverse transcriptase polymerase chain reaction (RT-PCR) assay that is capable of detecting low virus concentrations and a broad range of FCV isolates. The assay consisted of a 1-step RT-PCR reaction with primers delineating a 126-base-pair (bp) region ... More
Fluorescence imaging in human identity testing.
AuthorsWorley J, Lee S, Ma MS, Eisenberg A, Chen HY, Mansfield E
JournalBiotechniques
PubMed ID9232247
'We have investigated the use of fluorescence detection and the FluorImager S1 System (Molecular Dynamics) for analyzing a comprehensive set of human DNA typing tests. We used an alkaline phosphatase-conjugated YNH24 oligonucleotide probe to the repeat-containing D2S44 locus to detect both alleles in 50 ng of human genomic DNA (0.025 ... More
Visualization of mosaicism in tissues of normal and mismatch-repair-deficient mice carrying a microsatellite-containing transgene.
AuthorsHersh MN, Stambrook PJ, Stringer JR
JournalMutat Res
PubMed ID12175905
'To determine the frequency of mutation in different cell types of mammals, transgenic mice that allow mutant cells to be visualized in situ were used. These mice carry a defective allele of the human placental alkaline phosphatase (PLAP) gene. The allele does not produce enzyme because the reading frame is ... More
Angiotensin II stimulates the nuclear translocation of Smad2 and induces PAI-1 mRNA in rat hepatic stellate cells.
AuthorsKamada Y, Tamura S, Kiso S, Fukui K, Doi Y, Ito N, Terui Y, Saito T, Watanabe H, Togashi H, Kawata S, Matsuzawa Y
JournalHepatol Res
PubMed ID12697251
'It has recently been reported that angiotensin II (ANGII) stimulates gene expression of transforming growth factor-beta1 (TGF-beta1) and is involved in hepatic fibrosis. This effect is thought to be caused through ANGII type 1 receptor (ATR1). However, a role of ANGII at the postreceptor levels of TGF-beta1 signaling has not ... More
Direct quantification of fungal DNA from soil substrate using real-time PCR.
AuthorsFilion M, St-Arnaud M, Jabaji-Hare SH
JournalJ Microbiol Methods
PubMed ID12609725
'Detection and quantification of genomic DNA from two ecologically different fungi, the plant pathogen Fusarium solani f. sp. phaseoli and the arbuscular mycorrhizal fungus Glomus intraradices, was achieved from soil substrate. Specific primers targeting a 362-bp fragment from the SSU rRNA gene region of G. intraradices and a 562-bp fragment ... More
Selective detection of individual DNA molecules by capillary polymerase chain reaction.
AuthorsLi H, Xue G, Yeung ES
JournalAnal Chem
PubMed ID11321306
'On-line capillary polymerase chain reaction (PCR) coupled with laser-induced fluorescence detection was successfully demonstrated for individual DNA molecules. A single 30-microm-i.d. fused-silica capillary was used both as the reaction vessel and for isolating single molecules. SYBR green I dye was added into the reaction mixture for dynamic fluorescent labeling. Because ... More
Direct measurement of cell numbers in microtitre plate cultures using the fluorescent dye SYBR green I.
AuthorsMyers MA
JournalJ Immunol Methods
PubMed ID9671157
'The quantitation of cell numbers is essential for many experimental procedures. This study describes the use of the fluorescent DNA binding dye, SYBR green I, to quantitate accurately cell numbers in the wells of microtitre plates by determination of DNA content. The assay involves a single-step procedure and is sensitive ... More
Xer recombination in Escherichia coli. Site-specific DNA topoisomerase activity of the XerC and XerD recombinases.
AuthorsCornet F, Hallet B, Sherratt DJ
JournalJ Biol Chem
PubMed ID9268326
'Xer site-specific recombination functions in maintaining circular replicons in the monomeric state in Escherichia coli. Two recombinases of the bacteriophage lambda integrase family, XerC and XerD, are required for recombination at the chromosomal site, dif, and at a range of plasmid-borne sites. Xer recombination core sites contain the 11-base pair ... More
Enumeration of total bacteria and bacteria with genes for proteolytic activity in pure cultures and in environmental samples by quantitative PCR mediated amplification.
AuthorsBach HJ, Tomanova J, Schloter M, Munch JC
JournalJ Microbiol Methods
PubMed ID11869788
'Real-time quantitative PCR assays were developed for the absolute quantification of different groups of bacteria in pure cultures and in environmental samples. 16S rRNA genes were used as markers for eubacteria, and genes for extracellular peptidases were used as markers for potentially proteolytic bacteria. For the designed 16S rDNA TaqMan ... More
Improved mRNA quantitation in LightCycler RT-PCR.
AuthorsBall TB, Plummer FA, HayGlass KT
JournalInt Arch Allergy Immunol
PubMed ID12576739
'BACKGROUND: Real-time polymerase chain reaction (PCR) utilizing the LightCycler and similar systems is an increasingly used technique for quantitative reverse transcription (RT)-PCR of mRNA levels from genes of immunologic interest. A commonly encountered limitation with these systems is that the fluorescence induced by SYBR Green (a fluorophore that binds double-stranded ... More
Quantitative real-time reverse transcription-PCR assay for urokinase plasminogen activator, plasminogen activator inhibitor type 1, and tissue metalloproteinase inhibitor type 1 gene expressions in primary breast cancer.
AuthorsCastelló R, Estellés A, Vázquez C, Falcó C, España F, Almenar SM, Fuster C, Aznar J
JournalClin Chem
PubMed ID12142386
'BACKGROUND: The plasminogen activation system and matrix metalloproteinases (MMPs) play a key role in the degradation of basement membrane and extracellular matrix in tissue remodeling, cancer cell invasion, and metastasis. METHODS: Quantitative real-time reverse-transcription-PCR (RT-PCR) assays were developed to quantify urokinase-type plasminogen activator (uPA), plasminogen activator inhibitor type 1 (PAI-1), ... More
Position-specific expression of Hox genes along the gastrointestinal tract.
AuthorsYahagi N, Kosaki R, Ito T, Mitsuhashi T, Shimada H, Tomita M, Takahashi T, Kosaki K
JournalCongenit Anom (Kyoto)
PubMed ID15008896
'Hox genes play a critical role in morphogenesis of the early embryo along the anteroposterior axis. In mammals, 39 Hox genes with extensive homology are organized into 13 paralogous groups, forming four clusters on four separate chromosomes. The genes within each cluster are arranged in a 3'' to 5'' direction ... More
Mechanism of oligonucleotide release from cationic liposomes.
Authors Zelphati O; Szoka F C Jr;
JournalProc Natl Acad Sci U S A
PubMed ID8876163
'We propose a mechanism for oligonucleotide (ODN) release from cationic lipid complexes in cells that accounts for various observations on cationic lipid-nucleic acid-cell interactions. Fluorescent confocal microscopy of cells treated with rhodamine-labeled cationic liposome/ fluorescein-labeled ODN (F-ODN) complexes show the F-ODN separates from the lipid after internalization and enters the ... More
Effect of estrogen on the expression of matrix metalloproteinase (MMP)-1, MMP-3, and MMP-13 and tissue inhibitor of metalloproternase-1 in osteoarthritis chondrocytes.
AuthorsLee YJ, Lee EB, Kwon YE, Lee JJ, Cho WS, Kim HA, Song YW
JournalRheumatol Int
PubMed ID12684836
'The aim of this study was to evaluate the effect of estrogen on matrix metalloproteinase (MMP)-1, MMP-3, MMP-13, and tissue inhibitor of metalloproteinase (TIMP)-1 in osteoarthritic chondrocytes. Chondrocytes from the knee cartilage of 25 postmenopausal osteoarthritic (OA) patients were cultured under various conditions: 0 pg/mL, 50 pg/mL, 500 pg/mL, and ... More
High fidelity PCR with an off/on switch mediated by proofreading polymerases combining with phosphorothioate-modified primer.
AuthorsYang HL, Jiang HJ, Fang WY, Xu YY, Li K, Zhang J, Liao DF, He FC
JournalBiochem Biophys Res Commun
PubMed ID15670779
'In the initial report, introducing a single phosphorothioate modification at the very 3'' terminus of the oligodeoxynucleotide primer has been shown to effectively protect the oligodeoxynucleotide degradation due to the 3'' exonuclease activity. In this study, we reported a novel finding that phosphorothioate modification at the 3'' end of primers ... More
Rapid detection of apoptosis through real-time reverse transcriptase polymerase chain reaction measurement of the small cytoplasmic RNA Y1.
AuthorsAsselbergs FA, Widmer R
JournalAnal Biochem
PubMed ID12814625
'Apoptosis could be measured in mammalian cells by measuring the degradation of the small cytoplasmic human RNA Y1 (hY1) by real-time quantitative fluorescent reverse transcriptase polymerase chain reaction (RT-PCR). In FAS-antibody-treated Jurkat T cell leukemia cells degradation of hY1 occurred rapidly and was complete at about 6h. As in apoptotic ... More
Enumeration of viable anaerobic bacteria by solid phase cytometry under aerobic conditions.
AuthorsVermis K, Vandamme PA, Nelis HJ
JournalJ Microbiol Methods
PubMed ID11997163
'Classical enumeration methods for anaerobes are time-consuming and require special conditions. Solid phase cytometry (SPC) is a recent laser scanning technique for the quantitative detection of fluorescently labelled bacteria on a membrane filter that eliminates the need for a growth phase. Fluorescent labelling of cells results from the cleavage by ... More
Long Range Polymerase Chain Reaction-Based Diagnosis of Friedreich Ataxia in the Clinical Molecular Diagnostics Laboratory.
AuthorsWick MJ, Matthias-Hagen VL, Day JW, Gomez CM, Glennen RC
JournalMol Diagn
PubMed ID10096952
'Background: Friedreich ataxia, an autosomal recessive neurodegenerative disease, is one of the recently identified trinucleotide repeat disorders. Approximately 94% of affected individuals are homozygous for an intronic GAA repeat within the frataxin gene. The identification of this trinucleotide expansion as the causative mutation in the majority of affected individuals has ... More
The bile acid glycochenodeoxycholate induces trail-receptor 2/DR5 expression and apoptosis.
AuthorsHiguchi H, Bronk SF, Takikawa Y, Werneburg N, Takimoto R, El-Deiry W, Gores GJ
JournalJ Biol Chem
PubMed ID11507096
'Toxic bile salts induce hepatocyte apoptosis by both Fas-dependent and -independent mechanisms. In this study, we examined the cellular mechanisms responsible for Fas-independent, bile acid-mediated apoptosis. HuH-7 cells, which are known to be Fas deficient, were stably transfected with the sodium-dependent bile acid transporting polypeptide. The toxic bile acid glycochenodeoxycholate ... More
Deoxyadenosine analogs induce programmed cell death in chronic lymphocytic leukemia cells by damaging the DNA and by directly affecting the mitochondria.
AuthorsGenini D, Adachi S, Chao Q, Rose DW, Carrera CJ, Cottam HB, Carson DA, Leoni LM
JournalBlood
PubMed ID11071652
'Adenine deoxynucleosides induce apoptosis in quiescent lymphocytes and are thus useful drugs for the treatment of indolent lymphoproliferative diseases. To explain why deoxyadenosine and its analogs are toxic to a cell that is not undergoing replicative DNA synthesis, several mechanisms have been proposed, including the direct binding of dATP to ... More
The origin recognition complex marks a replication origin in the human TOP1 gene promoter.
'The locations of the origin recognition complex (ORC) in mammalian genomes have been elusive. We have therefore analyzed the DNA sequences associated with human ORC via in vivo cross-linking and chromatin immunoprecipitation. Antibodies specific for hOrc2 protein precipitate chromatin fragments that also contain other ORC proteins, suggesting that the proteins ... More
Effect of different extenders and storage temperatures on sperm viability of liquid ram semen.
AuthorsPaulenz H, Söderquist L, Pérez-Pé R, Berg KA
JournalTheriogenology
PubMed ID11991386
'Semen was collected with an artificial vagina from four adult rams. The ejaculates were pooled and diluted, using a split-sample technique, in four different extenders: one for milk (Mi), one for sodium citrate (Na), and two for Tris-based extenders (T1 and T2) including egg yolk. Thereafter, the diluted semen was ... More
Microarray identification of FMRP-associated brain mRNAs and altered mRNA translational profiles in fragile X syndrome.
AuthorsBrown V, Jin P, Ceman S, Darnell JC, O'Donnell WT, Tenenbaum SA, Jin X, Feng Y, Wilkinson KD, Keene JD, Darnell RB, Warren ST
JournalCell
PubMed ID11719188
'Fragile X syndrome results from the absence of the RNA binding FMR protein. Here, mRNA was coimmunoprecipitated with the FMRP ribonucleoprotein complex and used to interrogate microarrays. We identified 432 associated mRNAs from mouse brain. Quantitative RT-PCR confirmed some to be >60-fold enriched in the immunoprecipitant. In parallel studies, mRNAs ... More
Rhinovirus detection: comparison of real-time and conventional PCR.
AuthorsDagher H, Donninger H, Hutchinson P, Ghildyal R, Bardin P
JournalJ Virol Methods
PubMed ID15041207
'Rhinoviruses are important human respiratory viruses and the major causative agents of the common cold. Historically, detection of rhinovirus has been by virus culture and this was significantly improved by the use of PCR assays. Recently real-time PCR was developed but to date there have been no reported comparisons of ... More
Detection of bacterial contamination in starch and resin-based papermaking chemicals using fluorescence techniques.
AuthorsNohynek L, Saski E, Haikara A, Raaska L
JournalJ Ind Microbiol Biotechnol
PubMed ID12720090
'Rapid fluorescence techniques were evaluated for the detection of bacterial contaminants in papermaking chemicals including starch and the resin-based sizes and starch slurries used in the paper industry. Viable and non-viable bacterial cells were visualised by fluorescent probes and detected by epifluorescence microscopy and flow cytometry. The best discrimination ability ... More
Rapid diagnosis of alpha(o)-thalassemia using the relative quantitative PCR and the dissociation curve analysis.
AuthorsSangkitporn SK, Wangkahat K, Sangnoi A, Songkharm B, Charoenporn P, Sangkitporn S
JournalClin Lab Haematol
PubMed ID14641139
'A method for diagnosis of alpha(o)-thalassemia was developed based on detection of accumulated PCR product using SYBR Green I, a double-stranded DNA binding dye, and a fluorescence-detecting thermocycler. Primers were designed to specifically amplify - -SEA and - -Thai deletions of alpha(o)-thalassemia. Albumin was selected as the reference gene. The ... More
Quantitative approach to single-nucleotide polymorphism analysis using MALDI-TOF mass spectrometry.
AuthorsRoss P, Hall L, Haff LA
JournalBiotechniques
PubMed ID10997276
'Pooling of DNA samples before genotyping is a valuable means of streamlining large-scale genotyping efforts in disease association studies, single-nucleotide polymorphism (SNP) validation or mutant allele screening programs. In this report, we explore the application of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) to quantitative analysis of SNPs. The ... More
Distinct NFAT family proteins are involved in the nuclear NFAT-DNA binding complexes from human thymocyte subsets.
AuthorsAmasaki Y, Masuda ES, Imamura R, Arai K, Arai N
JournalJ Immunol
PubMed ID9498773
'The nuclear factor of activated T cells (NFAT) is involved in the transcriptional induction of cytokine and other immunoregulatory genes during an immune response. Among four distinct NFAT family members identified to date, mRNAs of NFAT1, NFATc, and NFATx are expressed in the thymus. Here, we report the distribution of ... More
[Real time PCR methodology for quantification of nucleic acids]
AuthorsTse C, Capeau J
JournalAnn Biol Clin (Paris)
PubMed ID12805005
'The polymerase chain reaction (PCR) has become an essential tool for molecular biologists and its introduction into nucleic acids detection systems has revolutionized the quantitative analysis of DNA and RNA. The technique has rapidly evolved over the last few years and the growing interest in quantitative applications of the PCR ... More
Two and three-color fluorescence flow cytometric analysis of immunoidentified viable bacteria.
AuthorsBarbesti S, Citterio S, Labra M, Baroni MD, Neri MG, Sgorbati S
JournalCytometry
PubMed ID10878564
'BACKGROUND: Traditional culture methods well established in the past and still in use are not able to detect the environmental microorganisms that exist in a viable but not culturable state. A number of different fluorescence-based assays have been developed over the past decade to detect and identify viable bacteria in ... More
Solution-based scanning for single-base alterations using a double-stranded DNA binding dye and fluorescence-melting profiles.
AuthorsElenitoba-Johnson KS, Bohling SD
JournalAm J Pathol
PubMed ID11549577
'DNA molecules differing by as little as a single-base substitution have traditionally been distinguished by gel electrophoresis-based methodologies that exploit differences in the sequence-specific properties of double-stranded DNA (dsDNA) such as melting temperature and secondary conformational configuration. By comparison, solution-based fluorescence methods using sequence-specific probes are limited to detecting mutations ... More
Rapid simultaneous amplification and detection of the MBR/JH chromosomal translocation by fluorescence melting curve analysis.
AuthorsBohling SD, King TC, Wittwer CT, Elenitoba-Johnson KS
JournalAm J Pathol
PubMed ID9916923
'Polymerase chain reaction (PCR) amplification and product analysis for the detection of chromosomal translocations, such as the t(14;18), has traditionally been a two-step process. PCR product detection has generally entailed gel electrophoresis and/or hybridization or sequencing for confirmation of assay specificity. Using a microvolume fluorimeter integrated with a thermal cycler ... More
Quantification of histone acetyltransferase and histone deacetylase transcripts during early bovine embryo development.
AuthorsMcGraw S, Robert C, Massicotte L, Sirard MA
JournalBiol Reprod
PubMed ID12533400
'Mammalian oocytes are very unique cells with an unlimited developmental potential. These totipotent cells are able to remove existing gene-expression patterns and to impose new ones. However, genome reprogramming is still a mystery. Posttranslational modifications by acetylation of the N-termini portion of histones composing the nucleosome are involved in genome ... More
Analysis of baculovirus aggregates using flow cytometry.
AuthorsJorio H, Tran R, Meghrous J, Bourget L, Kamen A
JournalJ Virol Methods
PubMed ID16364459
'Aggregation of viral particles represents a significant problem for baculoviral stock processing and storage. Aggregation may also affect the results of viral particle counting. A method using flow cytometry was previously developed in our lab to measure the concentration of baculovirus particles produced in insect cell cultures. In the present ... More
Rapid detection of subtelomeric deletion/duplication by novel real-time quantitative PCR using SYBR-green dye.
AuthorsBoehm D, Herold S, Kuechler A, Liehr T, Laccone F
JournalHum Mutat
PubMed ID15024731
'Telomeric chromosome rearrangements may cause mental retardation, congenital anomalies, miscarriages, and hematological malignancies. Automated detection of subtle deletions and duplications involving telomeres is essential for high-throughput screening procedures, but impractical when conventional cytogenetic methods are used. Novel real-time PCR quantitative genotyping of subtelomeric amplicons using SYBR-green dye allows high-resolution screening ... More
Quantification of the expression level of the gene encoding the catalytic subunit of telomerase in testicular tissue specimens predicts successful sperm recovery.
AuthorsSchrader M, Müller M, Schulze W, Heicappell R, Krause H, Straub B, Miller K
JournalHum Reprod
PubMed ID11756380
'BACKGROUND: The objective of the present study was to evaluate the quantitative detection of human telomerase reverse transcriptase (hTERT) mRNA as a new molecular diagnostic parameter in the work-up of testicular tissue specimens from patients presenting with non-obstructive azoospermia. M ETHODS: hTERT mRNA expression was quantified in 49 cryopreserved testicular ... More
Multiple-complete-digest restriction fragment mapping: generating sequence-ready maps for large-scale DNA sequencing.
AuthorsWong GK, Yu J, Thayer EC, Olson MV
JournalProc Natl Acad Sci U S A
PubMed ID9144219
'Multiple-complete-digest mapping is a DNA mapping technique based on complete-restriction-digest fingerprints of a set of clones that provides highly redundant coverage of the mapping target. The maps assembled from these fingerprints order both the clones and the restriction fragments. Maps are coordinated across three enzymes in the examples presented. Starting ... More
Sublethal injury and resuscitation of Candida albicans after amphotericin B treatment.
AuthorsLiao RS, Rennie RP, Talbot JA
JournalAntimicrob Agents Chemother
PubMed ID12654647
'Amphotericin B treatment was previously shown to inhibit Candida albicans reproduction and reduce the fluorescence of vitality-specific dyes without causing a corresponding increase in the fluorescence of the mortality-specific dyes bis-(1,3-dibutylbarbituric acid)trimethine oxonol and SYBR Green I. In the present study, we have confirmed these results and have shown that ... More
Quantitative analysis of kallikrein 15 gene expression in prostate tissue.
AuthorsStephan C, Yousef GM, Scorilas A, Jung K, Jung M, Kristiansen G, Hauptmann S, Bharaj BS, Nakamura T, Loening SA, Diamandis EP
JournalJ Urol
PubMed ID12478190
'PURPOSE: The newly discovered human kallikrein 15 gene KLK15 has been shown in preliminary analysis to be associated with more aggressive types of prostate cancer. We quantitatively measured and compared gene expression of KLK15 in malignant and benign prostate tissues. MATERIALS AND METHODS: Matched prostate tissue samples from the cancerous ... More
Quantitative detection of human immunodeficiency virus type 1 (HIV-1) proviral DNA in peripheral blood mononuclear cells by SYBR green real-time PCR technique.
AuthorsGibellini D, Vitone F, Schiavone P, Ponti C, La Placa M, Re MC
JournalJ Clin Virol
PubMed ID15018857
'BACKGROUND: The persistence of proviral human immunodeficiency virus type 1 (HIV-1) DNA reservoir represents one of the major drawbacks to the total eradication of HIV-1. The quantitative determination of proviral HIV-1 DNA load offers significant therapeutic information, especially when the HIV-1 RNA levels drop below the detectable limits during the ... More
Significance of size and nucleic acid content heterogeneity as measured by flow cytometry in natural planktonic bacteria.
AuthorsGasol JM, Zweifel UL, Peters F, Fuhrman JA, Hagström A
JournalAppl Environ Microbiol
PubMed ID10508078
'Total bacterial abundances estimated with different epifluorescence microscopy methods (4'',6-diamidino-2-phenylindole [DAPI], SYBR Green, and Live/Dead) and with flow cytometry (Syto13) showed good correspondence throughout two microcosm experiments with coastal Mediterranean water. In the Syto13-stained samples we could differentiate bacteria with apparent high DNA (HDNA) content and bacteria with apparent low ... More
Enumeration of 16S rDNA of Desulfotomaculum lineage 1 in rice field soil by real-time PCR with SybrGreen detection.
AuthorsStubner S
JournalJ Microbiol Methods
PubMed ID11997166
'Real-time PCR is a new and highly sensitive method for the quantification of microbial organisms in environmental samples. This work was conducted to evaluate real-time PCR with SybrGreen (SG) detection as quantification method for Desulfotomaculum lineage 1 organisms in samples of rice field soil. The method was optimized in several ... More
Molecular, biochemical and ecological characterisation of a bio-catalytic calcification reactor.
AuthorsHammes F, Boon N, Clement G, de Villiers J, Siciliano SD, Verstraete W
JournalAppl Microbiol Biotechnol
PubMed ID12883864
'Bio-catalytic calcification (BCC) reactors utilise microbial urea hydrolysis by autochthonous bacteria for the precipitation-removal of calcium, as calcite, from industrial wastewater. Due to the limited knowledge available concerning natural ureolytic microbial calcium carbonate (CaCO(3)) precipitation, the microbial ecology of BCC reactors has remained a black box to date. This paper ... More
Development of conventional and real-time PCR assays for detection of Legionella DNA in respiratory specimens.
AuthorsRantakokko-Jalava K, Jalava J
JournalJ Clin Microbiol
PubMed ID11474011
'The development and validation of a PCR assay based on the use of new 16S ribosomal DNA (rDNA)-targeted primers to detect Legionella DNA in respiratory specimens are described. The assay was originally developed as conventional PCR followed by electrophoretic detection and was then adapted to Lightcycler format with SYBR Green ... More
Versatile low-viscosity sieving matrices for nondenaturing DNA separations using capillary array electrophoresis.
AuthorsMadabhushi RS, Vainer M, Dolnik V, Enad S, Barker DL, Harris DW, Mansfield ES
JournalElectrophoresis
PubMed ID9059830
'The high-resolution separation of double-stranded DNA (dsDNA) has important applications in physical mapping strategies and in the analysis of polymerase chain reaction (PCR) products. Although high-resolution separations of dsDNA by capillary electrophoresis (CE) have been reported, pulsed fields were required to achieve complete resolution of DNA fragments beyond 23 kilobase ... More
Product differentiation by analysis of DNA melting curves during the polymerase chain reaction.
AuthorsRirie KM, Rasmussen RP, Wittwer CT
JournalAnal Biochem
PubMed ID9056205
'A microvolume fluorometer integrated with a thermal cycler was used to acquire DNA melting curves during polymerase chain reaction by fluorescence monitoring of the double-stranded DNA specific dye SYBR Green I. Plotting fluorescence as a function of temperature as the thermal cycler heats through the dissociation temperature of the product ... More