Amplification of human genomic DNA sequences with polymerase chain reaction using a single oligonucleotide primer

Background Dermatomycoses are a group of pathologic abnormalities frequently seen in clinical practice, and their prevalence has increased in recent decades. Diagnostic confirmation of mycotic infection in nails is essential because there are several pathologic conditions with similar clinical manifestations. The classical method for confirming the presence of fungus in nail is microbiological culture and the identification of morphological structures by microscopy. Methods We devised a nested polymerase chain reaction (PCR) that amplifies specific DNA sequences of dermatophyte fungus that is notably faster than the 3 to 4 weeks that the traditional procedure takes. We compared this new technique and the conventional plate culture method in 225 nail samples. The results were subjected to statistical analysis. Results We found concordance in 78.2% of the samples analyzed by the two methods and increased sensitivity when simultaneously using the two methods to analyze clinical samples. Now we can confirm the presence of dermatophyte fungus in most of the positive samples in just 24 hours, and we have to wait for the result of culture only in negative PCR cases. Conclusions Although this PCR cannot, at present, substitute for the traditional culture method in the detection of dermatophyte infection of the nails, it can be used as a complementary technique because its main advantage lies in the significant reduction of time used for diagnosis, in addition to higher sensitivity.

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Use of Multiple Displacement Amplification as Pre-polymerase Chain Reaction (Pre-PCR) to amplify genomic DNA of siphonapterids preserved for long periods in scientific collections

Parasites & Vectors ◽

10.1186/1756-3305-3-86 ◽

2010 ◽

Vol 3 (1) ◽

pp. 86 ◽

Cited By ~ 5

Author(s):

Daniel M Avelar ◽

Pedro M Linardi

Keyword(s):

Polymerase Chain Reaction ◽

Genomic Dna ◽

Multiple Displacement Amplification ◽

Chain Reaction ◽

Polymerase Chain ◽

Scientific Collections

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Polymerase chain reaction based mapping of rye involving repeated DNA sequences

Genome ◽

10.1139/g92-093 ◽

1992 ◽

Vol 35 (4) ◽

pp. 621-626 ◽

Cited By ~ 23

Author(s):

Peter M. Rogowsky ◽

Ken W. Shepherd ◽

Peter Langridge

Keyword(s):

Polymerase Chain Reaction ◽

Dna Sequences ◽

Chromosome 1 ◽

Repeated Dna ◽

Chain Reaction ◽

Pcr Marker ◽

Banding Patterns ◽

Repeated Dna Sequences ◽

Cereal Rye ◽

Polymerase Chain

A novel type of polymerase chain reaction (PCR) marker was developed for the mapping of cereal rye (Secale cereale). Primer pairs were synthesized targeting the insertion sites of three individual copies of the R173 family of rye specific repeated DNA sequences. While one primer was derived from a sequence within the respective R173 element, the second primer corresponded to a flanking region. The complex banding patterns obtained in rye allowed not only the mapping of the three R173 elements to certain chromosome regions of 1RS (the short arm of rye chromosome 1) but also the mapping of an additional 3–10 easily identifiable bands per primer pair to other rye chromosomes. Linkage mapping of a polymorphic 1R band derived from three rye cultivars demonstrated the presence of nonallelic, dominant markers in two independent crosses. Because of the high copy number of the R173 family (15 000 copies per diploid rye genome), its dispersion over the entire length of all chromosomes and the high number of markers obtained per primer pair, PCR markers based on the R173 family provide an almost unlimited source for well-spaced markers in rye mapping.Key words: polymerase chain reaction, mapping, repetitive DNA sequences, wheat, rye.

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Use of the polymerase chain reaction for detection of Fusarium graminearum in bulgur wheat

Food Science and Technology ◽

10.1590/s0101-20612012005000027 ◽

2012 ◽

Vol 32 (1) ◽

pp. 201-208 ◽

Cited By ~ 1

Author(s):

Carla Bertechini Faria ◽

Giovana Caputo Almeida-Ferreira ◽

Karina Bertechine Gagliardi ◽

Tatiane Cristina Albuquerque Alves ◽

Dauri José Tessmann ◽

...

Keyword(s):

Polymerase Chain Reaction ◽

Fusarium Graminearum ◽

Genomic Dna ◽

Solid Medium ◽

Food Contamination ◽

Chain Reaction ◽

Specific Pcr ◽

Culture Characteristics ◽

Mycotoxigenic Fungi ◽

Polymerase Chain

The detection of mycotoxigenic fungi in foodstuff is important because their presence may indicate the possible associated mycotoxin contamination. Fusarium graminearum is a wheat pathogen and a producer of micotoxins. The polymerase chain reaction (PCR) has been employed for the specific identification of F. graminearum. However, this methodology has not been commonly used for detection of F. graminearum in food. Thus, the objective of the present study was to develop a molecular methodology to detect F. graminearum in commercial samples of bulgur wheat. Two methods were tested. In the first method, a sample of this cereal was contaminated with F. graminearum mycelia. The genomic DNA was extracted from this mixture and used in a F. graminearum specific PCR reaction. The F. graminearum species was detected only in samples that were heavily contaminated. In the second method, samples of bulgur wheat were inoculated on a solid medium, and isolates having F. graminearum culture characteristics were obtained. The DNA extracted from these isolates was tested in F. graminearum specific PCR reactions. An isolate obtained had its trichothecene genotype identified by PCR. The established methodology could be used in surveys of food contamination with F. graminearum.

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