Evaluation of four methods for extraction of bacterial DNA from ruminal contents | Evaluación de cuatro métodos de extracción de DNA bacteriano de muestras de contenido ruminal
2009
Balmes, G., Institut de Recerca i Tecnologia Agroalimentàries, Caldes de Montbui, Barcelona (España). Remugants | Serrano, A., Institut de Recerca i Tecnologia Agroalimentàries, Caldes de Montbui, Barcelona (España). Remugants | Bach, A., Institut de Recerca i Tecnologia Agroalimentàries, Caldes de Montbui, Barcelona (España). Remugants | Arís, A., Institut de Recerca i Tecnologia Agroalimentàries, Caldes de Montbui, Barcelona (España). Remugants
The objective of this study was to compare four DNA extraction methods for the purification of microbial DNA from ruminal contents. The evaluated methods were 1) the RBB +C method which combines the cell lysis by bead beater with DNA extraction by silica columns, 2) the phenol-chloroform extraction method plus a cell lysis step by bead beater, 3) the Power Soil kit (MO BIO Laboratories, Inc) and 4) the Fast DNA SPIN Kit (Qbiogene, Inc.). We compared the efficiency in terms of quantity and quality of extracted DNA, which was used for RT-PCR quantification of four bacteria species in order to evaluate whether there was any bias in the DNA extraction yield depending on the microbial structural features. The results indicated that the DNA of Gram negative Selenomonas ruminantium and Prevotella ruminicola was extracted more efficiently with the Fast DNA SPIN Kit whereas the yield of extracted DNA of Gram positive Ruminococcus flavefaciens and Streptococcus bovis was higher with the phenol-chloroform method. Although in all tested methods the DNA extraction yield was related to the microbial structural properties, the phenol-chloroform method combined with bead beating gave acceptable yields for all the microorganisms evaluated in this study.
显示更多 [+] 显示较少 [-]