scholarly journals Optimisation of a multiplex PCR assay of nuclear microsatellite markers for population genetics and clone identification in Robinia pseudoacacia L.

2012 ◽  
Vol 61 (1-6) ◽  
pp. 142-148 ◽  
Author(s):  
H. Liesebach ◽  
E. Ewald
Keyword(s):  
Multiplex Pcr ◽  
Microsatellite Loci ◽  
Robinia Pseudoacacia ◽  
Cost Effective ◽  
Parentage Analysis ◽  
Paternity Analysis ◽  
Clone Identification ◽  
Multiplex Pcr Assay ◽  
Nuclear Microsatellite ◽  
Robinia Pseudoacacia L

AbstractBlack locust (Robinia pseudoacacia L.) is a tree species native to North America. The multipurpose tree is cultivated worldwide, but causes problems due to its partially invasive character. The application of nuclear microsatellite loci has many aims in population genetic studies. Here we introduce a very cost-effective method for combining the information of 14 nuclear microsatellite loci into two multiplex PCR sets as a contribution to greater standardisation and more comparable results. Combined non-exclusion probabilities for clone identification using example populations are estimated at between 1.37*E-5 and 1.67*E-11, and for paternity analysis for 1.59*E-4. The detected weak linkage between some microsatellite loci is not considered to be a substantial restriction to the reliability of the set of markers in providing an appropriate method for fingerprinting and parentage analysis.

scholarly journals Species-Specific Identification of Human Adenoviruses by a Multiplex PCR Assay

2000 ◽  
Vol 38 (11) ◽  
pp. 4114-4120 ◽  
Author(s):  
WanHong Xu ◽  
Mike C. McDonough ◽  
Dean D. Erdman
Keyword(s):  
Multiplex Pcr ◽  
Human Adenovirus ◽  
Cost Effective ◽  
Pcr Assay ◽  
Multiplex Pcr Assay ◽  
Amplicon Size ◽  
Species Specific ◽  
Fiber Gene ◽  
Amplification Reaction

A multiplex PCR assay was developed by using primers to the fiber gene that could differentiate human adenovirus (Ad) species A through F in a single amplification reaction. The assay correctly identified the species of all 49 recognized Ad prototype strains as well as 180 geographically and temporally diverse Ad field isolates. Ad serotype 6 (Ad6) (species C), Ad16 (species B), Ad31 (species A), and Ad40 and Ad41 (species F) could also be distinguished by amplicon size within each respective species. In comparison, a previously described Ad species-specific multiplex PCR assay that used primers to the Ad hexon gene gave equivocal results with several serotypes of species B, whereas our multiplex assay amplified all species B serotypes equally well. Our multiplex PCR assay will permit rapid, accurate, and cost-effective classification of Ad isolates.

scholarly journals Sequence-Selective Recognition of Extended-Spectrum β-Lactamase GES-2 by a Competitive, Peptide Nucleic Acid-Based Multiplex PCR Assay

2004 ◽  
Vol 48 (9) ◽  
pp. 3402-3406 ◽  
Author(s):  
Gerhard F. Weldhagen
Keyword(s):  
Nucleic Acid ◽  
Multiplex Pcr ◽  
Peptide Nucleic Acid ◽  
Large Scale ◽  
Cost Effective ◽  
Coding Region ◽  
Screening Programs ◽  
Extended Spectrum ◽  
Multiplex Pcr Assay ◽  
Novel Method

ABSTRACT Extended-spectrum β-lactamases (ESBLs) in Pseudomonas aeruginosa, such as GES-2, which compromises the efficacy of imipenem, tend to be geographically restricted. The CC-to-AA base pair substitution at positions 493 and 494 of the bla GES-2-coding region distinguishes this ESBL from bla GES-1 and the bla IBC-type genes, making it an ideal target for the development of a novel sequence-specific, peptide nucleic acid (PNA)-based multiplex PCR detection method. By using two primer pairs in conjunction with a PNA probe, this method provided an accurate means of identification of bla GES-2 compared to standard PCR and gene sequencing techniques when it was used to test 100 P. aeruginosa clinical isolates as well as previously published, well-described control strains encompassing all presently known genes in the bla GES-IBC ESBL family. This novel method has the potential to be used in large-scale, cost-effective screening programs for specific or geographically restricted ESBLs.

scholarly journals Single-Step Multiplex PCR Assay for Determining 92 Pneumococcal Serotypes

2016 ◽  
Vol 54 (8) ◽  
pp. 2197-2200 ◽  
Author(s):  
José M. Marimón ◽  
María Ercibengoa ◽  
Erica Santacatterina ◽  
Marta Alonso ◽  
Emilio Pérez-Trallero
Keyword(s):  
Capillary Electrophoresis ◽  
Multiplex Pcr ◽  
Disease Surveillance ◽  
Pneumococcal Disease ◽  
Cost Effective ◽  
Single Step ◽  
Pneumococcal Serotypes ◽  
Pcr Assay ◽  
Single Tube ◽  
Multiplex Pcr Assay

For pneumococcal disease surveillance, simple and cost-effective methods capable of determining all serotypes are needed. Combining a single-tube multiplex PCR with fluorescently labeled primers followed by amplicon analysis using automated fluorescent capillary electrophoresis, each serotype of 92 reference isolates and 297 recently collected clinical isolates was successfully determined.

A multiplex PCR assay for the simultaneous detection and discrimination of the sevenEimeriaspecies that infect domestic fowl

Parasitology ◽  
2003 ◽  
Vol 127 (4) ◽  
pp. 317-325 ◽  
Author(s):  
S. FERNANDEZ ◽  
A. H. PAGOTTO ◽  
M. M. FURTADO ◽  
Â. M. KATSUYAMA ◽  
A. M. B. N. MADEIRA ◽  
...  
Keyword(s):  
Multiplex Pcr ◽  
Domestic Fowl ◽  
Detection Threshold ◽  
Simultaneous Detection ◽  
Cost Effective ◽  
Pcr Assay ◽  
Multiplex Pcr Assay ◽  
Sensitivity Tests ◽  
Different Sources ◽  
Single Tube Reaction

This study reports the development of a novel multiplex PCR assay based on SCAR (Sequence-Characterised Amplified Region) markers for the simultaneous diagnosis of the 7Eimeriaspecies that infect domestic fowl. Primer pairs specific for each species were designed in order to generate a ladder of amplification products ranging from 200 to 811 bp. Sensitivity tests for each species were carried out, showing a detection threshold of 1–5 pg, which corresponds approximately to 2–8 sporulated oocysts. Distinct isolates of the 7Eimeriaspecies from different geographical sources were tested and successfully detected by the assay. All the species were amplified homogeneously, whether or not one of them was present in a high quantity, indicating that there was no cross-interference. The assay was also tested with different sources ofTaqDNA polymerase and thermocycler models, confirming the high reproducibility of the reaction. The economy of consumables and labour represented by a single-tube reaction greatly facilitates the molecular diagnosis of a large number of samples, making it appropriate for field epizootiological surveys. We propose the use of this multiplex PCR assay as a rapid and cost-effective diagnostic method for the detection and discrimination of the 7Eimeriaspecies that infect domestic fowl.

scholarly journals A Multiplex PCR for Simultaneous Detection of Three Zoonotic ParasitesAncylostoma ceylanicum,A. caninum,andGiardia lambliaAssemblage A

2015 ◽  
Vol 2015 ◽  
pp. 1-6 ◽  
Author(s):  
Wei Hu ◽  
Sheng Wu ◽  
Xingang Yu ◽  
Auwalu Yusuf Abullahi ◽  
Meiran Song ◽  
...  
Keyword(s):  
Multiplex Pcr ◽  
High Efficiency ◽  
Intestinal Parasites ◽  
Simultaneous Detection ◽  
Reaction System ◽  
Cost Effective ◽  
Multiplex Pcr Assay ◽  
Parasitic Zoonoses ◽  
Zoonotic Parasites ◽  
Sensitivity Specificity

Ancylostoma ceylanicum,A. caninum, andGiardia lambliaassemblage A are common intestinal parasites of dogs and cats; they can also infect humans, causing parasitic zoonoses. In this study, a multiplex PCR method was developed for simultaneous identification and detection of those three zoonotic parasites. Three pairs of specific primers were designed based on ITS sequence ofA. ceylanicumandA. caninumand TPI gene ofG. lambliaavailable in the GenBank. The multiplex PCR reaction system was established by optimizing the reaction condition, and a series of tests on the sensitivity, specificity, and clinical application were also conducted. Results showed that three target fragments were amplified specifically; the detection limit was 10 eggs for bothA. ceylanicumandA. caninum, 72 pg DNA forG. lamblia. Of 112 clinical fecal samples, 34.8% and 17.8% samples were positive forA. caninumandA. ceylanicum, respectively, while only 2.7% samples were positive forG. lambliaassemblage A. It is concluded that the established multiplex PCR assay is a convenient, rapid, cost-effective, and high-efficiency method for molecular detection and epidemiological investigation of three zoonotic parasites.

IgE-vermittelte Allergie gegen Robinienholz (Robinia pseudoacacia L.) – eine Fallbeschreibung

Pneumologie ◽  
2004 ◽  
Vol 58 (11) ◽  
Author(s):  
S Kespohl ◽  
R Merget ◽  
M Gellert ◽  
T Brüning ◽  
M Raulf-Heimsoth
Keyword(s):  
Robinia Pseudoacacia ◽  
Robinia Pseudoacacia L

Usefulness of a Multiplex PCR for Detection of Diarrheagenic Escherichia coli in a Diagnostic Microbiology Laboratory Setting

2016 ◽  
Vol 1 (2) ◽  
pp. 38-42 ◽  
Author(s):  
Khairun Nessa ◽  
Dilruba Ahmed ◽  
Johirul Islam ◽  
FM Lutful Kabir ◽  
M Anowar Hossain
Keyword(s):  
Escherichia Coli ◽  
Multiplex Pcr ◽  
Clinical Laboratory ◽  
Proteinase K ◽  
Microbiology Laboratory ◽  
Pcr Assay ◽  
E Coli ◽  
Diarrheagenic Escherichia Coli ◽  
Multiplex Pcr Assay ◽  
Diagnostic Microbiology

A multiplex PCR assay was evaluated for diagnosis of diarrheagenic Escherichia coli in stool samples of patients with diarrhoea submitted to a diagnostic microbiology laboratory. Two procedures of DNA template preparationproteinase K buffer method and the boiling method were evaluated to examine isolates of E. coli from 150 selected diarrhoeal cases. By proteinase K buffer method, 119 strains (79.3%) of E. coli were characterized to various categories by their genes that included 55.5% enteroaggregative E. coli (EAEC), 18.5% enterotoxigenic E. coli (ETEC), 1.7% enteropathogenic E. coli (EPEC), and 0.8% Shiga toxin-producing E. coli (STEC). Although boiling method was less time consuming (<24 hrs) and less costly (<8.0 US $/ per test) but was less efficient in typing E. coli compared to proteinase K method (41.3% vs. 79.3% ; p<0.001). The sensitivity and specificity of boiling method compared to proteinase K method was 48.7% and 87.1% while the positive and negative predictive value was 93.5% and 30.7%, respectively. The majority of pathogenic E. coli were detected in children (78.0%) under five years age with 53.3% under one year, and 68.7% of the children were male. Children under 5 years age were frequently infected with EAEC (71.6%) compared to ETEC (24.3%), EPEC (2.7%) and STEC (1.4%). The multiplex PCR assay could be effectively used as a rapid diagnostic tool for characterization of diarrheagenic E. coli using a single reaction tube in the clinical laboratory setting.Bangladesh J Med Microbiol 2007; 01 (02): 38-42

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