Comparison of Total DNA Extraction Methods for Microbial Community Form Polluted Soil

Tanase, Ana-Maria; Mereuță, Ioana; Chiciudean, Iulia; Ionescu, Robertina; Milea, Ligia; Cornea, Călina Petruţa; Vassu, Tatiana; Stoica, Ileana

doi:10.1016/j.aaspro.2015.08.102

Cited by 16 publications

(12 citation statements)

References 13 publications

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“…In this study, based on homologous sequence alignment method and clustering with information extracted from RDF and BLAST databases, the unique representative sequence reads were obtained, which were delimited for further analysis (as the Table 2 showed 21,230,11,424,30,526,25,666,16,686, and 14,032 effective reads of the CHC1, CHC2, CHC3, CHC4, CHC5 and CHC6). Using the software program Mothur and sequence identity criteria cut off of 97%, these sequences were assigned to some Operational taxonomic units (OTUs).…”

Section: Resultsmentioning

confidence: 99%

“…The deficiencies of traditional techniques are effectively offset since high-throughput sequencing techniques have been introduced [20]. The most fundamental step in molecular biology for microbial strains, and even more, for microbial community structure analysis is DNA extraction [21]. High-throughput sequencing technologies have a major impact on the life sciences by allowing the rapid accurate sequencing of very large numbers of about 200bp nucleotide sequences, in applications ranging from genome sequencing and resequencing to digital microarrays and ChIP-Seq experiments [22].…”

Section: Bioinformatics Data Analysismentioning

confidence: 99%

See 1 more Smart Citation

Study of Correlation between Milk Yield and Intestinal Microbes in Chinese Holstein Cows under the Same Diet

Xiuying¹,

Hongyu²,

Zhenya³

et al. 2018

J Microb Biochem Technol

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Section: Resultsmentioning

confidence: 99%

Section: Bioinformatics Data Analysismentioning

confidence: 99%

Study of Correlation between Milk Yield and Intestinal Microbes in Chinese Holstein Cows under the Same Diet

Xiuying¹,

Hongyu²,

Zhenya³

et al. 2018

J Microb Biochem Technol

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“…Several studies on this topic have been published recently. Most have analyzed only the quantity and quality of the DNA isolated by various methods (Dineen et al 2010;Knauth et al 2013;Mahmoudi et al 2011;Tanase et al 2015). Two studies demonstrated that different isolation methods and the use of different commercial kits can influence sequencing results and community analysis, but they focused on bacterial 16S rRNA genes (Bag et al 2016;Zielińska et al 2017).…”

Section: Introductionmentioning

confidence: 99%

Profiling soil microbial communities with next-generation sequencing: the influence of DNA kit selection and technician technical expertise

Soliman

Yang

Yamazaki

et al. 2017

Preprint

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Structure and diversity of microbial communities are an important research topic in biology, since microbes play essential roles in the ecology of various environments.Different DNA isolation protocols can lead to data bias and can affect results of nextgeneration sequencing. To evaluate the impact of protocols for DNA isolation from soil samples and also the influence of individual handling of samples, we compared results obtained by two researchers (R and T) using two different DNA extraction kits: (1) MO BIO PowerSoil ® DNA Isolation kit (MO_R and MO_T) and (2) NucleoSpin ® Soil kit (MN_R and MN_T). Samples were collected from six different sites on Okinawa Island, Japan. For all sites, differences in the results of microbial composition analyses (bacteria, archaea, fungi, and other eukaryotes), obtained by the two researchers using the two kits, were analyzed.For both researchers, the MN kit gave significantly higher yields of genomic DNA at all sites compared to the MO kit (ANOVA; P <0.006). In addition, operational taxonomic units for some phyla and classes were missed in some cases: Micrarchaea were detected only in the MN_T and MO_R analyses; the bacterial phylum Armatimonadetes was detected only in MO_R and MO_T; and WIM5 of the phylum Amoebozoa of eukaryotes was found only in the MO_T analysis. Our results suggest the possibility of handling bias; therefore, it is crucial that replicated DNA extraction be performed by at least two technicians for thorough microbial analyses and to obtain accurate estimates of microbial diversity.PeerJ Preprints | https://doi.org/10.7287/peerj.preprints.3073v1 | CC BY 4.0 Open Access |

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“…In this regard, it has been demonstrated that pretreatment of sediment samples to remove inhibitors and contaminants can be crucial. The application of common but appropriate reagents, such as EDTA or Tween-20, promotes their elimination for retrieval of total DNA from contaminated sediments (Fang et al, 2015), although the inclusion of glass beads is frequently used for their elimination (Tanase et al, 2015). One technique for purifying DNA with remnant humic acids is the electrodialysis method, which consists of the application of an electrical field to the DNA isolate, employing a dialysis membrane for specific extraction of DNA from the solution.…”

Section: Introductionmentioning

confidence: 99%

Research Article Inexpensive metagenomic DNA extraction protocol with high quality from marine sediments contaminated by petroleum hydrocarbons.

et al. 2017

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ABSTRACT. Marine environments are a reservoir of relevant information on dangerous contaminants such as hydrocarbons, as well as microbial communities with probable degradation skills. However, to access microbial diversity, it is necessary to obtain high-quality DNA. An inexpensive, reliable, and effective metagenomic DNA (mgDNA) extraction protocol from marine sediments contaminated with petroleum hydrocarbons was established in this study from modifications to Zhou's protocol. The optimization included pretreatment of sediment with saline solutions for the removal of contaminants, a second precipitation and enzymatic degradation of RNA, followed by purification of mgDNA extracted by electroelution. The results obtained indicated that the modifications applied to 12 sediments with total petroleum hydrocarbon (TPH) concentrations from 22.6-174.3 (µg/g dry sediment) yielded 20.3-321.3 ng/µL mgDNA with A 260 /A 280 and A 260 /A 230 ratios of 1.75 ± 0.08 and 1.19 ± 0.22, respectively. The 16S rRNA amplification confirmed the purity of the mgDNA. The suitability of this mgDNA extraction protocol lies in the fact that all chemical solutions utilized are common in all molecular biology laboratories, and the use of dialysis membrane does not require any sophisticated or expensive equipment, only an electrophoretic chamber.

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Comparison of Total DNA Extraction Methods for Microbial Community Form Polluted Soil

Cited by 16 publications

References 13 publications

Study of Correlation between Milk Yield and Intestinal Microbes in Chinese Holstein Cows under the Same Diet

Study of Correlation between Milk Yield and Intestinal Microbes in Chinese Holstein Cows under the Same Diet

Profiling soil microbial communities with next-generation sequencing: the influence of DNA kit selection and technician technical expertise

Research Article Inexpensive metagenomic DNA extraction protocol with high quality from marine sediments contaminated by petroleum hydrocarbons.

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