A simplified procedure to analyse mitochondrial DNA from industrial yeasts

López-Alonso, Victoria; Querol, Amparo; Ramón, Daniel; Fernández-Espinar, M. Teresa

doi:10.1016/s0168-1605(01)00483-4

Cited by 80 publications

(59 citation statements)

References 8 publications

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“…For COX1-PCR assays, the primers 3L (GCTTTAATTGGWGGWTTTGG), 3R (ATTGTCATACCATTTGTYCTYAT), 4L (GAAGTAGCAGGWGGWG GWGA), and 5R (GTTAGCTAAGGCWACWCCWGT) were used (27). PCR was performed in a 50-l volume containing 50 pmol of each primer, approximately 300 ng of DNA as a template, 0.25 mM concentrations of each deoxynucleoside triphosphate, 1.5 mM MgCl 2 , and 0.5 U of Taq polymerase.…”

Section: Methodsmentioning

confidence: 99%

“…Digestion of mtDNA with restriction enzymes such as HinfI or RsaI is associated with a high rate of polymorphism and has been used to study the authenticity of commercial wine yeast strains (14,40,41). A PCR method based on variations found on the mitochondrial COX1 (a gene coding for cytochrome oxidase) intron number and position has been validated to distinguish S. cerevisiae strains allowing researchers to monitor the evolution of wine fermentations conducted by commercial active dry yeast (27). Moreover, in many other studies, RAPD [random(ly) amplified polymorphic DNA]-PCR with different primers such as EL1 and LA1 has been successfully used to discriminate between wine yeast strains (2) and for differentiation of Saccharomyces species isolated in Brazil (18).…”

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confidence: 99%

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Biochemical and Molecular Characterization ofSaccharomyces cerevisiaeStrains Obtained from Sugar-Cane Juice Fermentations and Their Impact in Cachaça Production

Oliveira

Vicente

Fietto

et al. 2008

Appl Environ Microbiol

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Saccharomyces cerevisiae strains from different regions of Minas Gerais, Brazil, were isolated and characterized aiming at the selection of starter yeasts to be used in the production of cachaça, the Brazilian sugar cane spirit. The methodology established took into account the screening for biochemical traits desirable in a yeast cachaça producer, such as no H 2 S production, high tolerance to ethanol and high temperatures, high fermentative capacity, and the abilities to flocculate and to produce mycocins. Furthermore, the yeasts were exposed to drugs such as 5,5,5؆-trifluor-D,L-leucine and cerulenin to isolate those that potentially overproduce higher alcohols and esters. The utilization of a random amplified polymorphic DNA-PCR method with primers based on intron splicing sites flanking regions of the COX1 gene, as well as microsatellite analysis, was not sufficient to achieve good differentiation among selected strains. In contrast, karyotype analysis allowed a clear distinction among all strains. Two selected strains were experimentally evaluated as cachaça producers. The results suggest that the selection of strains as fermentation starters requires the combined use of biochemical and molecular criteria to ensure the isolation and identification of strains with potential characteristics to produce cachaça with a higher quality standard.Cachaça (pronounced "kha-sha-ssa"), the sugar cane spirit, is the most popular distilled beverage produced in Brazil. The annual production reaches 1.3 billion liters, with 15% being produced in more than 8,500 distilleries in the state of Minas Gerais.Traditional cachaça production relies on a spontaneous fermentative process that is mediated by the microbiota present in the cane juice wort and on the surface of equipments used in the productive process. It has been already demonstrated that in such systems there occurs a succession of yeasts, with Saccharomyces cerevisiae being the predominant species. Cachaça quality depends on the ecology of the microbial populations during an initial spontaneous fermentation (18,29,31,32,39). The fermentative process occurs through a continuously open fermentative process which is completed within 24 h and generally takes place from May to November, corresponding to the sugar cane harvesting period.Considering the conditions of production usually found in the cachaça distilleries, fermenting yeast populations have to face different types of stress (osmotic, high temperature, and high ethanol concentration). Besides, they might also present some characteristics such as a good fermentative power, no H 2 S production, killer activity, flocculation ability, and production of flavoring compounds. Taking all of these factors into account, we have developed a strategy to select yeast strains with appropriated characteristics to produce cachaça with potentially higher-quality standards (52). Parallel to the selection and development of S. cerevisiae strains toward ethanolic fermentations, molecular methods were developed and validated to study...

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Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

Biochemical and Molecular Characterization ofSaccharomyces cerevisiaeStrains Obtained from Sugar-Cane Juice Fermentations and Their Impact in Cachaça Production

Oliveira

Vicente

Fietto

et al. 2008

Appl Environ Microbiol

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“…Isolated strains were analysed by mitochondrial DNA restriction patterns (mtDNA RFLP) [3]. Strains with identical mtDNA RFLP patterns were grouped and one representative strain was further characterised by analysis of 10 S. cerevisiae specific microsatellite loci [4,5].…”

Section: Molecular Molecular Identification Identificationmentioning

confidence: 99%

Bioinformatic approaches for the genetic and phenotypic characterization of a Saccharomyces cerevisiae wine yeast collection

et al. 2008

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The objective of the present study was to compare genetic and phenotypic variation of 103 Saccharomyces cerevisiae strains isolated from winemaking environments. We used bioinformatics approaches to identify genetically similary strains with specific phenotypes and to estimate a strain's biotechnological potential. A S. cerevisiae collection, comprising 440 strains that were obtained from winemaking environments in Portugal has been constituted during the last years. All strains were genetically characterized by a set of eleven highly polymorphic microsatellites and showed unique allelic combinations. Using neural networks, a subset of 103 genetically most diverse strains was chosen for phenotypic analysis, that included growth in synthetic must media at various temperatures, utilization of carbon sources (glucose, ribose, arabinose, xylose, saccharose, galactose, rafinose, maltose, glycerol, potassium acetate and pyruvic acid), growth in ethanol containing media, evaluation of osmotic and oxidative stress resistance, H2S production and utilization of different nitrogen sources. Using supervised data mining approaches we have found that genotype represented with presence/absence of eleven microsatellites relates well with geographical location (performance evaluation using leave-out-out technique resulted in high performance scores; e.g., area under ROC curve was above 0.8 for a number of standard machine learning approaches tested). To find relations between phenotypes and genotypes, we used a two-step approach which first hierarchically clusters the strains according to their phenotype, and then tests if the resulting sub-clusters are identifiable using strain’s genetic data. Several groups of strains with similar phenotype profiles and common features in genotype were identified this way, and they are subject to further investigations. Financially supported by the programs POCI 2010 (FEDER/FCT, POCTI/AGR/56102/2004) and AGRO (ENOSAFE, Nº 762).

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“…A análise do perfil de restrição do DNA mitocondrial (mtDNA-RFLP) tem sido apontada como uma técnica eficiente para caracterizar e diferenciar linhagens industriais de S. cerevisiae (GuILLAMóN et al, 1996;LóPEz et al, 2001). Dessa forma, mtDNA-RFLP tem-se mostrado útil para diferenciar linhagens isoladas de um mesmo mosto e para monitorar a persistência e prevalência de linhagens específicas durante todo gerada na reação foi quantificada enzimaticamente com glicose oxidase e peroxidase, utilizando-se um kit enzimático comercial (Analisa Diagnóstica Ltda, Brasil).…”

Section: Introductionunclassified

Caracterização bioquímica e molecular de linhagens de Saccharomyces cerevisiae isoladas de fermentações artesanais de caldo de cana e melado para a produção de cachaça em Florianópolis-SC

Badotti¹,

Dário²,

Rosa³

et al. 2010

BJFT

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A simplified procedure to analyse mitochondrial DNA from industrial yeasts

Cited by 80 publications

References 8 publications

Biochemical and Molecular Characterization ofSaccharomyces cerevisiaeStrains Obtained from Sugar-Cane Juice Fermentations and Their Impact in Cachaça Production

Biochemical and Molecular Characterization ofSaccharomyces cerevisiaeStrains Obtained from Sugar-Cane Juice Fermentations and Their Impact in Cachaça Production

Bioinformatic approaches for the genetic and phenotypic characterization of a Saccharomyces cerevisiae wine yeast collection

Caracterização bioquímica e molecular de linhagens de Saccharomyces cerevisiae isoladas de fermentações artesanais de caldo de cana e melado para a produção de cachaça em Florianópolis-SC

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