Effect of DNA Extraction Method on the Apparent Microbial Diversity of Soil

Abstract

Four extraction methods, including a novel one, were compared for their efficiencies in producing DNA from three contrasting agricultural soils. Molecular analyses (PCR-denaturing gradient gel electrophoresis [DGGE] and clone libraries) focusing on different microbial groups were used as assessment criteria. Per soil, the DNA yields differed between extraction methods. Clear effects of method on apparent richness and community structure were found. Actinobacterial diversity based on soil DNA produced by two divergent methods revealed that a hitherto-undescribed group was obtained by the novel method.

Contemporary molecular tools in microbial ecology and their application to advancing biotechnology

“…The disadvantages of indirect methods are the longer processing time and lower DNA yields. One recent study demonstrated quantitatively the effect of DNA extraction methods on the microbial diversity of the sampled soil and concluded that the yield and diversity of DNA greatly depends on the extraction method and suggested that the use of a novel extraction method may result in having DNA from a microbial taxon that otherwise would not be identified (Inceoglu, Hoogwout et al 2010). …”

Technicalities and Glitches of Terminal Restriction Fragment Length Polymorphism (T-RFLP)

“…Past data on community profiling by T-RFLP and other methods revealed that use of a single method for DNA extraction can significantly underestimate the total number of bacterial ribotypes [23,25,26]. Several DNA extraction methods for diverse communities and habitats are available [19,20,23,25]. Therefore, a combination of different DNA extraction methods and the use of more than one genetic marker are recommended to obtain a more accurate evaluation of the diversity of the target organisms or functional groups [23,25,27,28].…”

“…Past data on community profiling by T-RFLP and other methods revealed that use of a single method for DNA extraction can significantly underestimate the total number of bacterial ribotypes [23,25,26]. Several DNA extraction methods for diverse communities and habitats are available [19,20,23,25].…”

“…Differential lysis of microbial cells, especially spore formers, during extraction of community DNA may lead to a bias in the relative amount of DNA in different samples [19][20][21][22][23][24][25]. Past data on community profiling by T-RFLP and other methods revealed that use of a single method for DNA extraction can significantly underestimate the total number of bacterial ribotypes [23,25,26].…”

“…Several DNA extraction methods for diverse communities and habitats are available [19,20,23,25]. Therefore, a combination of different DNA extraction methods and the use of more than one genetic marker are recommended to obtain a more accurate evaluation of the diversity of the target organisms or functional groups [23,25,27,28]. Multi-component amplification of the community DNA [28] and uneven distribution of copy number of the marker gene in community populations have their own issues [29].…”