Extraction and purification of DNA in rhizosphere soil samples for PCR-DGGE analysis of bacterial consortia

Method optimization for denaturing gradient gel electrophoresis (DGGE) analysis of microflora from Eucalyptus sp. wood chips intended for pulping

“…Some species remained unidentified due to poor visibility and close proximity of bands leading to underreporting of the full microbial spectrum. These challenges were also encountered by other authors (Gelsomino et al, 1999;Maarit-Niemi et al, 2001;Xue et al, 2008). Sample overloading and variation of DGGE gradients may resolve this issue.…”

“…Many of these studies relied on traditional culturing techniques to study microflora. Molecular methods relying on DNA or RNA extracted directly from environmental samples such as polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE), automated ribosomal intergenic spacer analysis (ARISA) and terminal restriction fragment length polymorphism (T-RFLP) are increasingly being used, particularly in analyzing community structure of microorganisms from different environments, for example, soil (Gelsomino et al, 1999;Maarit-Niemi et al, 2001;Hoshino and Matsumoto, 2007;Xue et al, 2008), compost (Adams and Frostick, 2009;Novinscak et al, 2009), wastewater (Moura et al, 2009;Liu et al, 2010;Chen et al, 2013), food (Greaves, 1975;Miller et al, 1999;Ji et al, 2004;Handschur et al, 2005) and decaying chip piles intended for fuel (Rajala et al, 2010;Rajala et al, 2011). Several recent reports focused on understanding the microbial effects on storage of woody biomass intended for biofuel production, as well as composting (Ashraf et al, 2007;Adams and Frostick, 2009;Novinscak et al, 2009).…”

Soil microbial community analysis in the environmental risk assessment of transgenic plants

“…However, the recovery of DNA/RNA from soil has not yet become a routinely practiced work, and often requires refinement of the extraction and purification conditions due to the extreme diversity of the physical, chemical and biological properties of soils. It is also well recognized that the methods employed for DNA or RNA extraction can also bias the subsequent results of microbial community analyses, in terms of both qualitative and quantitative interpretation of data (Maarit-Niemi et al 2001).…”

“…In addition, it is known that some bacterial groups have variations in length and sequence between intragenomic copies of ribosomal operons (intercistronic heterogeneity), for both the RNA gene coding and intergenic regions (Maarit-Niemi et al 2001). Furthermore, different species will have different gene copy numbers and this can also bias the interpretation of fingerprinting results (Liu et al 1997).…”

Molecular Analysis of Bacterial Community DNA in Sludge Undergoing Autothermal Thermophilic Aerobic Digestion (ATAD): Pitfalls and Improved Methodology to Enhance Diversity Recovery

“…A range of commercial extraction kits are now available and many are attractive in terms of short operation times, ease of performance, lack of refrigeration necessary in the field, and the minimal set of chemicals and equipment required. Indeed there have been numerous reports of their successful application in a variety of niches [30,31]. However the composition of ingredients is often proprietary, a full understanding of the protocol and what is being achieved is often absent and most importantly the extent to which the full biodiversity of the niche under examination is being explored is rarely determined or examined.…”