estandarización de un protocolo sencillo para la extracción de adn genómico de levaduras

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ID: 147911
2009
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Abstract
Se estandarizó un protocolo rápido, sencillo y de bajo costo para la extracción de ADN genómico de levaduras a partir de lisis de la pared celular mediante tratamiento enzimático y precipitación por alcoholes. El empleo de la enzima Beta-glucoronidasa en reemplazo de la enzima Zimolasa, permitió obtener ADN en alta concentración (124,9±30,2 ng/μl) y de buena calidad (A260/A280 nm =1,86±0,1), ideal para su uso en estudios de biología molecular. Además, se adicionó un paso de incubación del ADN obtenido a 100° C para inactivar ADNasas. La calidad del ADN obtenido fue evaluada por medio de la amplificación de la región ITS1-5.8S-ITS2, presentando bandas definidas y cuantificables (entre 380 y 880 pb) ideales para estudios de identificación molecular y filogenia. Abstract: A quick, simple and low-cost protocol for extracting genomic DNA from yeast by cell wall lysis involving enzymatic treatment and alcoholic precipitation was standardised. Higher DNA yields (124.9±30.2 ng/μl) were obtained by using beta-glucuronidase instead of zymolyase; these had very high quality (A260/A280 nm = 1.86±0.1) and would be suitable for use in molecular biology assays. Moreover, a DNAse inactivation step was also introduced by incubation at 100 °C to further ensure DNA stability. DNA quality was assayed by PCR amplification of the ITS1-5.8S-ITS2 region, revealing defined, quantifiable 380 to 880 bp bands. These results show that the protocol is ideal for molecular identification and phylogenetic studies.
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osorio-cadavid2009revistaestandarizacin Use this key to autocite in the manuscript while using SciMatic Manuscript Manager or Thesis Manager
Authors Esteban Osorio-Cadavid, Mauricio Ramírez, William Andrés López, Luz Adriana Mambuscay
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Journal gigascience
Year 2009
DOI
DOI not found
URL
https://revistas.unal.edu.co/index.php/biotecnologia/article/view/10339
Keywords
yeast dna extraction

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