Enhancement of soil DNA extraction by the use of a hand held mixer

Costa, J. N. da; Oliveira, Virgínia Carla de

doi:10.1590/s1517-83822003000400004

Cited by 2 publications

(4 citation statements)

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“…The efficiency of soil DNA extraction obtained with a mini mixer (Moulinex, Brazil) was seen before by [12], but they did not amplify their samples to be analysed by PCR-DGGE technique and now the present work comes with additional information about this method because this was compared with a commercial kit that is greatly used to direct DNA extraction from bulk soil to study bacterial communities.…”

Section: Statistical Analysis Of Denaturing Gradient Gel Electrophore...mentioning

confidence: 81%

“…Extraction of cells from soil samples were done following a modification of the method proposed by [12] that used a hand held mini mixer to induce cell lyses in substitution to a bead beater which is more expensive. Four grams of soil were transferred to a sterilized glass beaker containing 4 mL of 0.12 mol -1 phosphate buffer (pH 8.0).…”

Section: Extraction Of Cells From Soil and Dna Extractionmentioning

confidence: 99%

“…These can turn the laboratorial routines faster and more tangible. A method of soil DNA extraction using a hand held mini mixer that costs 1% of a bead beater and that yielded higher amounts of pure DNA was developed [12], but the PCR products were not analysed by DGGE (Denaturing Gradient Gel Electrophoresis).…”

Section: Introductionmentioning

confidence: 99%

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Comparison of two DNA Extraction Protocols Representing Bacterial Community from Bulk Soil by PCR-DGGE

Martins

Passos

Xavier

et al. 2015

BMRJ

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Aims: The objective of this work was to investigate the efficiency of DNA extraction from bulk soil using the Britânia® mini mixer combined with phenol extraction to study bacterial communities, from three sampling sites localized in São Gonçalo-RJ, Brazil, comparing it to a commercial kit by the PCR-DGGE technique. Study Design: Molecular fingerprints of bacterial communities in bulk soil from three sampling sites were generated by DGGE after 16S rDNA gene amplification. Place and Duration of Study: Faculdade de Formação de Professores-UERJ and Embrapa Agrobiologia between April 2010 and April 2013. Methodology: Samples of DNA, in triplicate, extracted from bulk soil at three sampling sites localized in São Gonçalo-RJ, Brazil, were obtained by two protocols of DNA extraction. The DNA Original Research Articlesamples were used as template to amplify the 16S rDNA gene using specific primers to α and β-Proteobacteria groups. The PCR products were used for a second amplification using the primers F968GC and R1401 for subsequent DGGE analysis. The products from the second amplification were subjected to DGGE and band patterns were statistically analysed. Results: The band profiles obtained from the protocol that used a hand held mini mixer were intense and showed higher similarity among the triplicates from the sampling sites. Although both methods were capable of achieving a representative DNA profile from the soil bacterial community, the band patterns produced by them for the same areas were different. Conclusion:The DGGE profile of specific groups such as α-and β-Proteobacteria was a useful tool to compare the two soil DNA extraction protocols and also to compare the community structure of the different sampled areas. The DNA extraction protocol that used the Britânia® mini mixer produced band profiles with higher values of richness, but missed some bacterial targets as the commercial kit did. Both protocols have validity for the study of bacterial communities in bulk soil. The clusters of band profiles obtained via 16S rDNA PCR-DGGE indicated differences in bacterial communities of bulk soil from the three sampling sites for different ecological succession stages localized in São Gonçalo, RJ. The diversity analysis showed that the α-Proteobacteria group was predominant in bulk soil from these sites.

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Section: Statistical Analysis Of Denaturing Gradient Gel Electrophore...mentioning

confidence: 81%

Section: Extraction Of Cells From Soil and Dna Extractionmentioning

confidence: 99%

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Comparison of two DNA Extraction Protocols Representing Bacterial Community from Bulk Soil by PCR-DGGE

Martins

Passos

Xavier

et al. 2015

BMRJ

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“…In cases where the method was used multiple times in a single study, each method was counted once per study to avoid artificially inflating the method's frequency of use SDS sodium dodecyl sulfate, CTAB cetyltrimethylammonium bromide, PVPP polyvinyl polypyrrolidone a Performed by either hand squeezing or sieving b Centrifugation procedure denotes differential centrifugation was performed for cell separation (Chen et al 2003a, b), 14 (Chen et al 2003a, b), 15 (Costa and de Oliveira 2003), 16 (Fus et al 2003), 17 (Gabor et al 2003), 18 (Uz et al 2003), 19 (Webster et al 2003), 20 (Grubb and Dehority 1976), 21 (Miller et al 1986), 22 , 23 , 24 , 25 (Olubobokun et al 1987), 26 (Barsuhn et al 1988), 27 (Barlaz et al 1989a, b), 28 (Whitehouse et al 1994), 29 (Martin-Orue et al 1998, 30 (Yanagita et al 2000), 31 (Courtois et al 2001), 32 (Bockelmann et al 2003), 33 (Ranilla and Carro 2003), 34 (Frostegard et al 1999), 35 (Tien et al 1999), 36 (Luna et al 2006), 37 (Wikstrom et al 1996), 38 (Zhou et al 1996), 39 (Miller et al 1999), 40 (Volossiouk et al 1995), 41 (Krause et al 2001), 42 (Ranjard et al 1998), 43 (Sharma et al 2003), 44 (Cheng et al 1991), 45 (Huang et al 2003), 46 (Huang et al 2004), 47 (Fortin et al 2004), 48 (Minato and Suto 1978), 49 (Kudo et al 1987), 50 (Rasmussen et al 1989), 51 (Minato and Suto 1981) Springs (NC) Transfer Station and appeared consistent with refuse composition given by the USEPA (2005). The r...…”

Section: Methodsmentioning

confidence: 99%

Critical evaluation of solid waste sample processing for DNA-based microbial community analysis

et al. 2010

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Landfills represent a unique microbial ecosystem and play a significant role in global biogeochemical processes. The study of complex ecosystems such as landfills using DNA-based techniques can be advantageous since they allow for analysis of uncultured organisms and offer higher resolution in measuring demographic and metabolic (functional) diversity. However, sample acquisition and processing from refuse is challenging due to material heterogeneity. Decomposed refuse was used to evaluate the effect of seven sample processing methods on Bacteria and Archaea community structure using T-RFLP. Bias was assessed using measured richness and by comparing community structure using multi-dimensional scaling (MDS). Generally, direct methods were found to be most biased while indirect methods (i.e., removal of cellular material from the refuse matrix before DNA extraction) were least biased. An indirect method using PO₄ buffer gave consistently high bacterial and archaeal richness and also resulted in 28 and 34% recovery of R. albus and M. formicicum spiked into refuse, respectively. However, the highest recovery of less abundant T-RFs was achieved using multiple processing methods. Results indicate differences in measured T-RF diversity from studies of landfill ecosystems could be caused by methodological (i.e., processing method) variation rather than refuse heterogeneity or true divergence in community structure.

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Enhancement of soil DNA extraction by the use of a hand held mixer

Abstract: The efficiency of soil DNA extraction using a bead beater homogenizer (Biospec Products -Germany) was compared to that obtained with a hand held mixer (Moulinex -Brazil). The hand held mixer costs 100 times less and extracted seven times more crude DNA than the bead beater.

Cited by 2 publications

References 5 publications

Comparison of two DNA Extraction Protocols Representing Bacterial Community from Bulk Soil by PCR-DGGE

Comparison of two DNA Extraction Protocols Representing Bacterial Community from Bulk Soil by PCR-DGGE

Critical evaluation of solid waste sample processing for DNA-based microbial community analysis

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