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Retrotransposon microsatellite amplified polymorphism strain fingerprinting markers applicable to various mushroom species.

Le QV, Won HK, Lee TS, Lee CY, Lee HS, Ro HS - Mycobiology (2008)

Bottom Line: To utilize marY1 as a molecular marker for the DNA fingerprinting of mushrooms, oligonucleotides marY1-LTR-L and marY1-LTR-R were designed on the basis of highly conserved regions from the multiple sequence alignment of 30 marY1 sequences retrieved from a nucleotide sequence database.In accordance with Retrotransposon Microsatellite Amplified Polymorphism (REMAP) fingerprinting methodology, the two oligonucleotides were utilized together with the short sequence repeat primers UBC807 and UBC818 for polymerase chain reaction using templates from different mushroom genomic DNAs.Among the tested oligonucleotides, the marY1-LTR-L and UBC807 primer set yielded the greatest amount of abundance and variation in terms of DNA band numbers and patterns.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Research Institute of Life Sciences, Gyeongsang National University, Chinju 660-701, Korea.

ABSTRACT
The retrotransposon marY1 is a gypsy family retroelement, which is detected ubiquitously within the fungal taxonomic groups in which mushrooms are included. To utilize marY1 as a molecular marker for the DNA fingerprinting of mushrooms, oligonucleotides marY1-LTR-L and marY1-LTR-R were designed on the basis of highly conserved regions from the multiple sequence alignment of 30 marY1 sequences retrieved from a nucleotide sequence database. In accordance with Retrotransposon Microsatellite Amplified Polymorphism (REMAP) fingerprinting methodology, the two oligonucleotides were utilized together with the short sequence repeat primers UBC807 and UBC818 for polymerase chain reaction using templates from different mushroom genomic DNAs. Among the tested oligonucleotides, the marY1-LTR-L and UBC807 primer set yielded the greatest amount of abundance and variation in terms of DNA band numbers and patterns. This method was successfully applied to 10 mushroom species, and the primer set successfully discriminated between different commercial mushroom cultivars of the same strains of 14 Pleurotus ostreatus and 16 P. eryngii. REMAP reproducibility was superior to other popular DNA fingerprinting methodologies including the random amplified polymorphic DNA method.

No MeSH data available.


Related in: MedlinePlus

Comparison of the reproducibility of REMAP to RAPD. Three mushroom genomic DNA samples (S1, S2, S3) and three commercial thermostable DNA polymerases (1: i-starTaq; 2: Taq polymerase; 3: Ex Taq) were used to assess the reproducibility of RAPD (A) and REMAP (B). PCR mixtures and conditions were optimal for each enzyme, according to the manufacturers' instructions.
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Figure 3: Comparison of the reproducibility of REMAP to RAPD. Three mushroom genomic DNA samples (S1, S2, S3) and three commercial thermostable DNA polymerases (1: i-starTaq; 2: Taq polymerase; 3: Ex Taq) were used to assess the reproducibility of RAPD (A) and REMAP (B). PCR mixtures and conditions were optimal for each enzyme, according to the manufacturers' instructions.

Mentions: The genetic analysis conducted on the fungal genus Fellomyces previously showed that RAPD and amplified fragment length polymorphism (AFLP) demonstrate a higher potential in distinguishing strains within species than do other DNA sequence-based methods (Lopandic et al., 2005). In particular, the RAPD method was applied successfully in the verification of P. eryngii strains from various hosts from a wide range of geographical origins (Zervakis et al., 2001). However, there are concerns about the reproducibility of this PCR-based technique that have been discussed elsewhere (Skroch and Nienhuis, 1995; Jones et al., 1997). It has been widely recognized that PCR mixture and reaction conditions are factors upon which the reproducibility of this PCR-based technique is solely based. To assess the reproducibility of our Com3-based REMAP, we evaluated the DNA band patterns from both the RAPD and REMAP analyses using different thermostable DNA polymerases. Three genomic DNA samples of P. ostreatus were employed as templates for the RAPD and REMAP analyses with three different commercial DNA polymerases (Fig. 3).


Retrotransposon microsatellite amplified polymorphism strain fingerprinting markers applicable to various mushroom species.

Le QV, Won HK, Lee TS, Lee CY, Lee HS, Ro HS - Mycobiology (2008)

Comparison of the reproducibility of REMAP to RAPD. Three mushroom genomic DNA samples (S1, S2, S3) and three commercial thermostable DNA polymerases (1: i-starTaq; 2: Taq polymerase; 3: Ex Taq) were used to assess the reproducibility of RAPD (A) and REMAP (B). PCR mixtures and conditions were optimal for each enzyme, according to the manufacturers' instructions.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3755187&req=5

Figure 3: Comparison of the reproducibility of REMAP to RAPD. Three mushroom genomic DNA samples (S1, S2, S3) and three commercial thermostable DNA polymerases (1: i-starTaq; 2: Taq polymerase; 3: Ex Taq) were used to assess the reproducibility of RAPD (A) and REMAP (B). PCR mixtures and conditions were optimal for each enzyme, according to the manufacturers' instructions.
Mentions: The genetic analysis conducted on the fungal genus Fellomyces previously showed that RAPD and amplified fragment length polymorphism (AFLP) demonstrate a higher potential in distinguishing strains within species than do other DNA sequence-based methods (Lopandic et al., 2005). In particular, the RAPD method was applied successfully in the verification of P. eryngii strains from various hosts from a wide range of geographical origins (Zervakis et al., 2001). However, there are concerns about the reproducibility of this PCR-based technique that have been discussed elsewhere (Skroch and Nienhuis, 1995; Jones et al., 1997). It has been widely recognized that PCR mixture and reaction conditions are factors upon which the reproducibility of this PCR-based technique is solely based. To assess the reproducibility of our Com3-based REMAP, we evaluated the DNA band patterns from both the RAPD and REMAP analyses using different thermostable DNA polymerases. Three genomic DNA samples of P. ostreatus were employed as templates for the RAPD and REMAP analyses with three different commercial DNA polymerases (Fig. 3).

Bottom Line: To utilize marY1 as a molecular marker for the DNA fingerprinting of mushrooms, oligonucleotides marY1-LTR-L and marY1-LTR-R were designed on the basis of highly conserved regions from the multiple sequence alignment of 30 marY1 sequences retrieved from a nucleotide sequence database.In accordance with Retrotransposon Microsatellite Amplified Polymorphism (REMAP) fingerprinting methodology, the two oligonucleotides were utilized together with the short sequence repeat primers UBC807 and UBC818 for polymerase chain reaction using templates from different mushroom genomic DNAs.Among the tested oligonucleotides, the marY1-LTR-L and UBC807 primer set yielded the greatest amount of abundance and variation in terms of DNA band numbers and patterns.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Research Institute of Life Sciences, Gyeongsang National University, Chinju 660-701, Korea.

ABSTRACT
The retrotransposon marY1 is a gypsy family retroelement, which is detected ubiquitously within the fungal taxonomic groups in which mushrooms are included. To utilize marY1 as a molecular marker for the DNA fingerprinting of mushrooms, oligonucleotides marY1-LTR-L and marY1-LTR-R were designed on the basis of highly conserved regions from the multiple sequence alignment of 30 marY1 sequences retrieved from a nucleotide sequence database. In accordance with Retrotransposon Microsatellite Amplified Polymorphism (REMAP) fingerprinting methodology, the two oligonucleotides were utilized together with the short sequence repeat primers UBC807 and UBC818 for polymerase chain reaction using templates from different mushroom genomic DNAs. Among the tested oligonucleotides, the marY1-LTR-L and UBC807 primer set yielded the greatest amount of abundance and variation in terms of DNA band numbers and patterns. This method was successfully applied to 10 mushroom species, and the primer set successfully discriminated between different commercial mushroom cultivars of the same strains of 14 Pleurotus ostreatus and 16 P. eryngii. REMAP reproducibility was superior to other popular DNA fingerprinting methodologies including the random amplified polymorphic DNA method.

No MeSH data available.


Related in: MedlinePlus