scholarly journals Βελτιστοποίηση της τεχνικής RAPD - PCR (random amplified polymorphic DNA-PCR) και εφαρμογή της στη μελέτη απομονώσεων Leishmania του σκύλου στην Ελλάδα

2000 ◽  
Author(s):  
Αναστασία Διάκου
Keyword(s):  
Random Amplified Polymorphic Dna ◽  
Rapd Pcr ◽  
Dna Pcr

scholarly journals Characterization of verocytotoxin-producingEscherichia coliO157 isolates from patients with haemolytic uraemic syndrome in Western Europe

1995 ◽  
Vol 115 (1) ◽  
pp. 1-3 ◽  
Author(s):  
A. E. Heuvelink ◽  
N. C. A. J. van de Kar ◽  
J. F. G. M. Meis ◽  
L. A. H. Monnens ◽  
W. J. G. Melchers
Keyword(s):  
Haemolytic Uraemic Syndrome ◽  
Western Europe ◽  
Random Amplified Polymorphic Dna ◽  
E Coli ◽  
Pcr Fingerprinting ◽  
Phage Types ◽  
Rapd Pcr ◽  
Typing Methods ◽  
Dna Pcr

SummaryFifty verocytotoxin (VT)-producingEscherichia coli(VTEC) strains of serogroup O157 were characterized by phage typing, polymerase chain reaction (PCR) for VT genes and theE. coliattaching and effacing (eae) gene, and random amplified polymorphic DNA–PCR (RAPD–PCR) fingerprinting. The collection represented isolates obtained from patients with diarrhoea-associated haemolytic-uraemic syndrome (D+ HUS) and their family contacts, isolated in the Netherlands, Belgium and Germany between 1989 and 1993. Based on isolates from separate families (n= 27) seven different phage types were identified, types 2 (44%) and 4 (33%) were predominant. Eighty-five percent of the strains contained only VT2 gene sequences and 15% both VT1 and VT2. All strains of the dominant phage types 2 and 4 carried the VT2 gene. Strains that belonged to the minor phage types 8, 14, 32 carried both VT1 and VT2 genes, with the exception of two isolates identified as phage types 49 and 54 which contained only VT2 genes. All O157 VTEC strains possessed the chromosomally-locatedeaegene, which indicates its usefulness as virulence marker. RAPD–PCR fingerprinting identified four distinct banding patterns, with one profile found among 79% of the strains. Based on the combined results of all typing methods used in this study, the collection of 50 O157 VTEC strains could be divided into nine distinct groups. Strains isolated from different persons within one family could not be distinguished by any of these methods. The data suggest that O157 VTEC strains are members of one clone that has become widely distributed.

Differentiation of faecal Escherichia coli from human and animal sources by random amplified polymorphic DNA-PCR (RAPD-PCR)

2004 ◽  
Vol 50 (1) ◽  
pp. 193-198 ◽  
Author(s):  
D. Venieri ◽  
A. Vantarakis ◽  
G. Komninou ◽  
M. Papapetropoulou
Keyword(s):  
Escherichia Coli ◽  
Rapd Analysis ◽  
Random Amplified Polymorphic Dna ◽  
Hierarchical Cluster ◽  
Group Method ◽  
Randomly Amplified Polymorphic Dna ◽  
Arbitrary Primers ◽  
E Coli ◽  
Rapd Pcr ◽  
Dna Pcr

In this study the assessment of randomly amplified polymorphic DNA (RAPD) analysis was established as a molecular epidemiological tool. RAPD analysis was performed to differentiate faecal Escherichia coli isolates from human and animal sources. E. coli strains (128) were isolated from human and animal faeces (from cattle and sheep). Genomic DNA was extracted and randomly amplified polymorphic DNA-PCR (RAPD-PCR) fingerprinting was performed. Seven arbitrary primers were tested with a view to discriminating between E. coli isolates from humans and E. coli isolates from animals. RAPD profiles were analysed with hierarchical cluster analysis using an unweighted pair group method. RAPD profiles obtained with three of the tested primers (1247, 1290 and 1254) established a distinct differentiation between E. coli isolates from humans and E. coli from animals. Low levels of misclassification and high levels of specificity make RAPD a sensitive, efficient and reliable means of distinguishing closely related strains.

scholarly journals Characteristics of the Genetic Variation of a Swamp Buffalo (Bubalusbubalis) of South Sumatra Based on Polymerase Chain Reaction-Random Amplified Polymorphic DNA (PCR-RAPD)

2018 ◽  
Vol 68 ◽  
pp. 01002
Author(s):  
Yuanita Windusari ◽  
Laila Hanum ◽  
Arum Setiawan ◽  
Veronika Larasati
Keyword(s):  
Genetic Variation ◽  
Random Amplified Polymorphic Dna ◽  
Molecular Approach ◽  
Chain Reaction ◽  
Rapd Pcr ◽  
Polymerase Chain ◽  
Polymorphic Bands ◽  
Swamp Buffalo ◽  
Dna Pcr ◽  
Cluster 2

Swamp buffalo (Bubalusbubalis) is one of the endemic species that become a wealth of genetic resources of South Sumatra. This study aims to the genetic variation and relationships of kinship 6 variants of swamp buffalo South Sumatera. The methods used by the molecular approach using RAPD-PCR primer 5 i.e. ILO 1204, ILO 1212, ILO 525, OPW 03 and OPY 13. Data was analyzed using SPSS ver 16.0 and presented in dendrogram. The results of the amplification, all primary produce band with a total of 63 band of DNA (14.92%) with an average of every primary produce 12.6 band of DNA. The most primary produce DNA polymorphic bands namely OPW 03 (23.81%) and ILO 1204 (20.63%), while the primary ILO 525 (0.00%) do not generate polymorphic bands. Genetic variation of swamp buffalo has a low genetic variation with 14.92% percentage it generated polymorphic bands. The results of the dendogram obtained two clusters namely cluster 1 included Kerbau Tanduk Bulat, Kerbau Tanduk Langit, Kerbau Tanduk Melintang and Kerbau Tanduk Dungkul, while the cluster 2 of them Kerbau Bule and Kerbau Rebah Belakang. Swamp buffalo variants that have the closest genetic distance. Kerbau Tanduk Langit and Kerbau Tanduk Bulat with 856 coefficient similarity, while the farthest Kerbau Tanduk Langit and Kerbau Bule with the coefficient similarity -972. Swamp buffalo (Bubalusbubalis) of South Sumatera, which consists of 6 variants of buffalo have low genetic variation and inbreeding of closekinship.

scholarly journals Utility of Random Amplified Polymorphic DNA PCR and TaqMan Automated Detection in Molecular Identification ofAspergillus fumigatus

1998 ◽  
Vol 36 (7) ◽  
pp. 2057-2062 ◽  
Author(s):  
Mary E. Brandt ◽  
Arvind A. Padhye ◽  
Leonard W. Mayer ◽  
Brian P. Holloway
Keyword(s):  
Detection System ◽  
Random Amplified Polymorphic Dna ◽  
Taqman Assay ◽  
Pcr Cloning ◽  
System A ◽  
Fungal Isolates ◽  
Rapd Pcr ◽  
Rapd Primer ◽  
Species Specific ◽  
Dna Pcr

We developed a method for the identification of Aspergillus fumigatus fungal isolates by using random amplified polymorphic DNA (RAPD) PCR (RAPD-PCR) cloning and the TaqMan LS50B fluorogenic detection system (Perkin-Elmer Corp., Applied Biosystems, Foster City, Calif.). DNA from seven clinically important Aspergillusspecies was screened by RAPD-PCR to identify section- or species-specific amplicons. With the OPZ19 RAPD primer a 1,264-bp product was amplified from all A. fumigatus strains initially examined but not from other species. A partial DNA sequence of this product was used to design a specific primer pair, which generated a single 864-bp fragment with DNA from 90 of 100 A. fumigatus isolates when a “touchdown” (65→55°C) annealing protocol was used. The TaqMan system, a fluorogenic assay which uses the 5′→3′ endonuclease activity of Taq DNA polymerase, detected this 864-bp product with DNA from 89 of these 90 A. fumigatus strains; 1 DNA sample generated an indeterminate result. With DNA from three morphologically typical A. fumigatus isolates, six white (“albino”) A. fumigatus isolates, and five of six Neosartoryaspecies (non-A. fumigatus members of the sectionFumigati), the 864-bp product was amplified differentially at an annealing temperature of 56°C but not with the touchdown annealing format. No amplicon was detected with DNA from 56 isolates of heterologous Aspergillus, Penicillium, andPaecilomyces species or from Neosartorya fennelliae; TaqMan assay results were either negative (51 isolates) or indeterminate (5 isolates) for all isolates. This RAPD-PCR and TaqMan assay offers promise as a nucleic acid-based system that can be used for the identification of filamentous fungal isolates and that requires no postamplification sample manipulations.

scholarly journals Oenological Potential of Autochthonous Saccharomyces cerevisiae Yeast Strains from the Greek Varieties of Agiorgitiko and Moschofilero

Beverages ◽  
2021 ◽  
Vol 7 (2) ◽  
pp. 27
Author(s):  
Dimitrios Kontogiannatos ◽  
Vicky Troianou ◽  
Maria Dimopoulou ◽  
Polydefkis Hatzopoulos ◽  
Yorgos Kotseridis
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Alcoholic Fermentation ◽  
Ethanol Tolerance ◽  
Random Amplified Polymorphic Dna ◽  
Yeast Strains ◽  
Rapd Pcr ◽  
Autochthonous Yeast ◽  
Polymerase Chain ◽  
Grape Varieties ◽  
H2s Production

Nemea and Mantinia are famous wine regions in Greece known for two indigenous grape varieties, Agiorgitiko and Moschofilero, which produce high quality PDO wines. In the present study, indigenous Saccharomyces cerevisiae yeast strains were isolated and identified from spontaneous alcoholic fermentation of Agiorgitiko and Moschofilero musts in order to evaluate their oenological potential. Random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) recovered the presence of five distinct profiles from a total of 430 yeast isolates. The five obtained strains were evaluated at microvinifications trials and tested for basic oenological and biochemical parameters including sulphur dioxide and ethanol tolerance as well as H2S production in sterile grape must. The selected autochthonous yeast strains named, Soi2 (Agiorgitiko wine) and L2M (Moschofilero wine), were evaluated also in industrial (4000L) fermentations to assess their sensorial and oenological characteristics. The volatile compounds of the produced wines were determined by GC-FID. Our results demonstrated the feasibility of using Soi2 and L2M strains in industrial fermentations for Agiorgitiko and Moschofilero grape musts, respectively.

scholarly journals Antibiotic Susceptibility and Genotype Patterns of Escherichia coli, Klebsiella pneumonia and Pseudomonas aeruginosa Isolated from Urinary Tract Infected Patients

2010 ◽  
Vol 59 (3) ◽  
pp. 207-212 ◽  
Author(s):  
M.I. ABOU-DOBARA ◽  
M.A. DEYAB ◽  
E.M. ELSAWY ◽  
H.H. MOHAMED
Keyword(s):  
Escherichia Coli ◽  
Pseudomonas Aeruginosa ◽  
Urinary Tract ◽  
Antibiotic Susceptibility ◽  
Susceptibility Testing ◽  
Random Amplified Polymorphic Dna ◽  
Test Methods ◽  
Klebsiella Pneumonia ◽  
E Coli ◽  
Rapd Pcr

Thirty nine isolates of Escherichia coli, twenty two isolates of Klebsiella pneumoniae and sixteen isolates of Pseudomonas aeruginosa isolated from urinary tract infected patients were analyzed by antimicrobial susceptibility typing and random amplified polymorphic DNA (RAPD)-PCR. Antibiotic susceptibility testing was carried out by microdilution and E Test methods. From the antibiotic susceptibility, ten patterns were recorded (four for E. coli, three for K. pneumoniae and three for P. aeruginosa respectively). Furthermore, genotyping showed seventeen RAPD patterns (seven for E. coli, five for K. pneumoniae and five for P. aeruginosa respectively). In this study, differentiation of strains of E. coli, K. pneumoniae and P. aeruginosa from nosocomial infection was possible with the use of RAPD.

scholarly journals Genetic basis for clarithromycin resistance among isolates of Mycobacterium chelonae and Mycobacterium abscessus.

1996 ◽  
Vol 40 (7) ◽  
pp. 1676-1681 ◽  
Author(s):  
R J Wallace ◽  
A Meier ◽  
B A Brown ◽  
Y Zhang ◽  
P Sander ◽  
...  
Keyword(s):  
Point Mutation ◽  
Random Amplified Polymorphic Dna ◽  
Mycobacterium Abscessus ◽  
Single Step ◽  
23S Rrna ◽  
Mycobacterium Chelonae ◽  
Clarithromycin Resistance ◽  
Gene Coding ◽  
Disseminated Disease ◽  
Dna Pcr

Resistance to clarithromycin among isolates of Mycobacterium chelonae and M. abscessus was observed in 18 of 800 (2.3%) patients tested between 1990 and 1995. Patients whose isolates were resistant had either disseminated disease or chronic lung disease, and the resistant isolates were recovered after clarithromycin monotherapy. Sequencing of the gene coding for the 23S rRNA peptidyltransferase region revealed a point mutation involving adenine at position 2058 (38%) or adenine at position 2059 (62%) in 20 of 20 relapse isolates from the first 13 patients identified. By pulsed-field gel electrophoresis or random amplified polymorphic DNA PCR, initial and relapse isolates were shown to have identical DNA patterns. M. chelonae and M. abscessus isolates were found to have only a single chromosomal copy of the rRNA operon, thus making them susceptible to single-step mutations. Thus, clarithromycin resistance in these species of rapidly growing mycobacteria relates to a point mutation in the gene coding for 23S rRNA and occurs in limited clinical situations, but was identified in almost 5% of isolates tested in 1995.

scholarly journals Interlaboratory Study on Thermal Cycler Performance in Controlled PCR and Random Amplified Polymorphic DNA Analyses

2001 ◽  
Vol 47 (1) ◽  
pp. 47-55 ◽  
Author(s):  
Ginny C Saunders ◽  
Juliet Dukes ◽  
Helen C Parkes ◽  
Johanne H Cornett
Keyword(s):  
Random Amplified Polymorphic Dna ◽  
Positive Sample ◽  
Interlaboratory Study ◽  
Pcr Assay ◽  
Single Product ◽  
The United Kingdom ◽  
Rapd Pcr ◽  
Block Temperature ◽  
Thermal Cycler ◽  
Correct Target

Abstract Background: Intercomparisons of PCR-based data between laboratories require an assurance of assay reproducibility. We performed an interlaboratory study to investigate the contribution made by a variety of thermal cyclers to PCR performance as measured by interblock reproducibility and intrablock repeatability. Methods: Two standardized assays designed to minimize the introduction of non-thermal-cycler-dependent variations were evaluated by 18 laboratories in the United Kingdom, using 33 thermal cyclers of various makes and models. We used a single-product (590 bp) PCR, established in our laboratory as a robust and specific reaction. The second reaction, a multiproduct random amplified polymorphic DNA (RAPD) PCR, was known to be more susceptible to small changes in block temperature and was therefore considered a way of assessing block uniformity with respect to temperature. Assay repeatability data were analyzed with respect to temperature calibration status, the type of temperature control mechanism, thermal cycler age, and the presence of oil overlay or heated lid systems. Results: All (100%) of the laboratories produced the correct target for the single-product PCR assay, although substantial variation in yield in replicate reactions was observed in 9.4% of these. The RAPD reaction generated results that varied extensively both within the same block and between different thermal cyclers. For eight replicates of a positive sample, 88% intrablock repeatability was demonstrated in calibrated thermal cyclers, which decreased to 63% in noncalibrated instruments. Conclusions: Irrespective of the make and model of thermal cycler, temperature-calibrated instruments consistently generated more repeatable RAPD data than noncalibrated instruments. Guidelines are offered on optimizing and monitoring thermal cycler performance.

Sign in / Sign up

Export Citation Format

Share Document