Direct colony polymerase chain reaction (PCR): An efficient technique to rapidly identify and distinguish Mycosphaerella fijiensis and Mycosphaerella musicola

2012 ◽  
Vol 11 (33) ◽  
Author(s):  
Roberto Vázquez-Euán
Keyword(s):  
Polymerase Chain Reaction ◽  
Efficient Technique ◽  
Mycosphaerella Fijiensis ◽  
Chain Reaction ◽  
Colony Polymerase Chain Reaction ◽  
Mycosphaerella Musicola ◽  
Polymerase Chain

scholarly journals Colony Polymerase Chain Reaction withSchizosaccharomyces pombe

2016 ◽  
Vol 2016 (5) ◽  
pp. pdb.prot090993 ◽  
Author(s):  
Johanne M. Murray ◽  
Adam T. Watson ◽  
Antony M. Carr
Keyword(s):  
Polymerase Chain Reaction ◽  
Chain Reaction ◽  
Colony Polymerase Chain Reaction ◽  
Polymerase Chain

scholarly journals Mycosphaerella fijiensis, Causal Agent of Black Sigatoka of Musa spp. Found in Puerto Rico and Identified by Polymerase Chain Reaction

Plant Disease ◽  
2006 ◽  
Vol 90 (5) ◽  
pp. 684-684 ◽  
Author(s):  
B. M. Irish ◽  
R. Goenaga ◽  
R. C. Ploetz
Keyword(s):  
Polymerase Chain Reaction ◽  
Puerto Rico ◽  
Production Costs ◽  
Natural Resource Institute ◽  
Mycosphaerella Fijiensis ◽  
Black Sigatoka ◽  
Chain Reaction ◽  
Potato Dextrose Broth ◽  
Musa Spp ◽  
Polymerase Chain

Black Sigatoka, also known as black leaf streak, is caused by Mycosphaerella fijiensis Morelet (anamorph Pseudocercospora fijiensis (Morelet) Deighton). It is the most significant disease of bananas and plantains (Musa spp.) because most of the economically important cultivars of exported and staple commodities are highly susceptible. The Caribbean is one of the few regions of the world where black Sigatoka is not widespread. Black Sigatoka has been reported in the Bahamas, Cuba, Hispaniola, and Jamaica (2). Yellow Sigatoka, caused by M. musicola Leach (anamorph P. musae (Zimm.) Deighton), has been recognized in Puerto Rico since 1938-1939 (3). In August 2004, symptoms resembling black Sigatoka were first observed in Añasco, Puerto Rico by extension personnel from the University of Puerto Rico. Since black and yellow Sigatoka produce similar disease symptoms, a survey was conducted in the western banana- and plantain-production region of Puerto Rico to confirm the presence of black Sigatoka. Leaf samples were collected from production fields near the towns of Las Marias, Maricao, and Añasco. Single-ascospore isolates were recovered using the discharge technique from moistened pseudothecia in necrotic lesions that were inverted over water agar, and ascospores were transferred to potato dextrose agar. The isolates were subcultured in potato dextrose broth for mycelium production. DNA was isolated from mycelium with the FastDNA kit (Q-Biogen, Irvine, CA) for 19 isolates. Internal transcribed spacer as well as the 5.8s rDNA regions were polymerase chain reaction amplified with primers specific to M. fijiensis or M. musicola (1). Amplification products (˜1,100 bp) were observed for 18 of the 19 isolates, 6 of which were M. fijiensis and the remaining 12 were M. musicola, while the positive controls for both species were also amplified with the respective primer pairs. M. fijiensis was recovered from production fields close to all three towns. The source of M. fijiensis in Puerto Rico is unclear, but it may have originated from introduced leaf material and/or wind dispersed ascospores from neighboring countries. The presence of black Sigatoka in Puerto Rico will most likely increase production costs where fungicide applications will be needed to maintain yields. The USDA-ARS, Tropical Agriculture Research Station is the official Musa spp. germplasm repository for the National Plant Germplasm System. As such, efforts are underway to introduce and evaluate black Sigatoka disease-resistant clones that can satisfy local and export market criteria. References: (1) A. Johnasen. Detection of Sigatoka leaf spot pathogens of banana by the polymerase chain reaction. Chatman, UK, Natural Resource Institute, 1997. (2) R. C. Ploetz. Plant Dis. 88:772, 2004. (3) R. H. Stover. Trop. Agric. Trinidad. 39:327, 1962.

High performance Nanogold™-in situ hybridzation and its use in the detection of hybridized and PCR-amplified microscopical preparations

1996 ◽  
Vol 54 ◽  
pp. 896-897
Author(s):  
G. W. Hacker ◽  
I. Zehbe ◽  
J. Hainfeld ◽  
A.-H. Graf ◽  
C. Hauser-Kronberger ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Messenger Rna ◽  
High Performance ◽  
Detection System ◽  
Direct Methods ◽  
Genetic Alterations ◽  
Chain Reaction ◽  
Protein Encoding ◽  
Polymerase Chain

In situ hybridization (ISH) with biotin-labeled probes is increasingly used in histology, histopathology and molecular biology, to detect genetic nucleic acid sequences of interest, such as viruses, genetic alterations and peptide-/protein-encoding messenger RNA (mRNA). In situ polymerase chain reaction (PCR) (PCR in situ hybridization = PISH) and the new in situ self-sustained sequence replication-based amplification (3SR) method even allow the detection of single copies of DNA or RNA in cytological and histological material. However, there is a number of considerable problems with the in situ PCR methods available today: False positives due to mis-priming of DNA breakdown products contained in several types of cells causing non-specific incorporation of label in direct methods, and re-diffusion artefacts of amplicons into previously negative cells have been observed. To avoid these problems, super-sensitive ISH procedures can be used, and it is well known that the sensitivity and outcome of these methods partially depend on the detection system used.

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