A Simple Silica-based Method for Metagenomic DNA Extraction from Soil and Sediments

Rojas-Herrera, Rafael; Narváez-Zapata, José Alberto; Zamudio-Maya, Marcela; Mena-Martínez, M E

doi:10.1007/s12033-008-9061-8

Cited by 51 publications

(29 citation statements)

References 18 publications

(6 reference statements)

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…The precipitated nanocrystals adsorbed proteins (Figure 3) via interaction between the protein group (NH 4 + ) and the crystal group (PO 4 -) or complex formation between HA-Ca 2 + --OOC-protein keeping only the nucleic acids of the ground material in contact with organic phases of the extraction process. The hydroxyapatite and octacalcic phosphate precipitates formed the cell debris, based on the principle underlying the use of silica beads for DNA (Rojas-Herrera et al, 2008); this idea is already used in HPLC (Guastaldi and Aparecida, 2010). Formation of these insoluble phosphates is similar to the biomimicking process proposed by Abe et al (1990).…”

Section: Resultsmentioning

confidence: 99%

New efficient DNA extraction method to access the microbiome of Ricinus communis seeds

Santos

Dias²,

Amaral³

et al. 2013

Genet. Mol. Res.

View full text Add to dashboard Cite

ABSTRACT. Ricinus communis (castor bean) seeds are used to produce an alcohol-soluble oil that is used in more than 400 industrial processes. Despite its economic importance, there has been little research on the endophytic microbiota of castor bean seeds. This microbiota is important for plant metabolic processes and may have considerable biotechnological potential, such as production of lipases and plant growth promoter agents. We evaluated several DNA extraction methodologies in order to access the microbial diversity of castor bean through a metagenomic approach. Based on our observations, we developed a new methodology that takes advantage of the low solubility of calcium phosphates and the high affinity of these phosphates for proteins and polysaccharides. The extracted DNA quality was evaluated by PCR, using a selective primer pair for bacterial and mitochondrial 16S rDNA genes (799F and 1492R). We found this methodology quantitatively and qualitatively more efficient than the other approaches. In evaluating this new extraction methodology, we found that the difficulties of DNA extraction from castor bean seeds, such as abundant oil, polysaccharides, phenolic compounds, and plant enzymes, could be overcome. The resulting extracts had high concentration and purity, ©FUNPEC-RP www.funpecrp.com.br Genetics and Molecular Research 12 (3): 3128-3135 (2013) New efficient DNA extraction to access oil seed microbiome 3129 and they were obtained faster than with previous methods. The samples contained virtually all of the DNA, including the microbial DNA; this was validated by PCR analysis.

show abstract

Section: Resultsmentioning

confidence: 99%

New efficient DNA extraction method to access the microbiome of Ricinus communis seeds

Santos

Dias²,

Amaral³

et al. 2013

Genet. Mol. Res.

View full text Add to dashboard Cite

show abstract

“…Successful results were obtained with protocols 4, 5 and 6. Protocol 4, described by Rojas-Herrera et al (2008), uses lysozyme and glass milk, which raises the price of DNA extraction; however, this procedure is also fast and in our hands has been successful for other types of samples, e.g., soil, sediments and marine water; thus, we decided to use it for comparison. This protocol was capable of extracting DNA suitable for PCR amplification (Figure 1, lane 1).…”

Section: Discussionmentioning

confidence: 99%

“…Increased understanding of microbial communities in MFC is necessary to design and control the system effectively. To this end, molecular techniques have gained preference for culture-independent bacterial identification (Rojas-Herrera et al, 2008;Ryu et al, 2011). However, the capability of determining microbial diversity largely depends on the quality of the DNA in question.…”

Section: Introductionmentioning

confidence: 99%

“…There is a plethora of protocols for DNA extraction of microbial communities (Zhou et al, 1996;Krsek and Wellington, 1999;Griffiths et al, 2000;Wechter et al, 2003;Chaudhuri et al, 2006;Rojas-Herrera et al, 2008;Ning et al, 2009), including homemade protocols and commercial kits. The latter often allow rapid recovery of nucleic acids, although some kits fail in obtaining nucleic acids in some samples (Hoshino and Matsumoto, 2005;Luna et al, 2006;Hou et al, 2010).…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Methodology Simple and inexpensive DNA extraction protocol for studying the bacterial composition of sludges used in microbial fuel cells

Canto-Canché¹,

Tzec-Simá²,

Vázquez-Loría³

et al. 2013

Genet. Mol. Res.

Self Cite

View full text Add to dashboard Cite

ABSTRACT. Bacteria oxidize organic matter and nutrients to produce electric energy in microbial fuel cells (MFC) -a technology of increasing importance because of its sustainability. To improve the performance of MFCs, it is necessary not only to gain a better understanding of MFC engineering designs, but also to improve the understanding of the composition of the microbial communities in MFCs. Fast and efficient DNA extraction protocols that are suitable for extracting diverse bacterial genomes are necessary to identify the bacterial diversity present in MFCs and to further monitor the dynamic changes of microbial communities. This study focused on testing different direct cell lysis protocols to extract DNA from a microbial sludge harvested DNA extraction of bacterial consortium in MFC from an MFC. The protocol that achieved the best results was based on a previous study, but was modified by eliminating a chaotropic salt and the special columns used for nucleic acid purification. The efficiency of this less expensive and more straightforward protocol was confirmed by PCR amplification of the 16S rRNA gene and denaturing gradient gel electrophoresis analysis, which confirmed the extraction of multiple genomes. The sequences of 10 clones revealed the presence of phyla, Proteobacteria, Firmicutes and Actinobacteria, comprising both Gramnegative and Gram-positive bacteria. Some of these bacteria were identified at the genus level, e.g., Clostridium, Pseudoxanthomonas, Tistrella, and Enterobacter; these genera have been described in active sludges from wastewater treatment, supporting the congruency of our results. Therefore, this protocol is a useful tool for analysis of the bacteria responsible for energy production in MFCs.

show abstract

“…Para la extracción de ADN se pesaron 1.5 g de suelo seco de cada muestra y se colocaron en tubos con 3.5 mL de NaH 2 PO 4 (0.1 M) y lisozima (10 mg/mL, pH 4.5), bajo agitación (100 rpm), durante 1 h. De cada tubo se tomaron tres alícuotas (1 mL) y se trataron de acuerdo con el procedimiento descrito por Rojas-Herrera et al (2008). Los extractos de ADN así obtenidos, se visualizaron por electroforesis en agarosa (1 %) con bromuro de etidio (10 mg/mL) como agente de tinción.…”

Section: Extracción De Adn Microbiano Del Suelounclassified

Dinámica De Poblaciones Bacterianas Y Actividad Deshidrogenasa Durante La Biorremediación De Suelo Recién Contaminado E Intemperizado Con Hidrocarburos

Ortíz-Maya¹,

Escalante-Espinosa²,

Ramírez-Saad

et al. 2017

Rev. Int. Contam. Ambie.

View full text Add to dashboard Cite

Palabras clave: actividad enzimática, DGGE, restauración de suelos, secuenciación RESUMEN Se evaluó la actividad deshidrogenasa y la dinámica de poblaciones bacterianas nativas en suelo contaminado con hidrocarburos durante su biorremediación por bioestimulación y atenuación natural. Se realizaron ensayos a escala mesocosmos, en la que se compararon dos condiciones: 1) contaminación reciente (28 días) y 2) intemperización (entre 28 a 56 días), con suelo artificialmente contaminado (30 000 mg de hidrocarburos totales de petróleo /kg suelo). Se estimó la riqueza de las comunidades bacterianas con base en los perfiles genómicos generados mediante electroforesis en gel con gradiente de desnaturalizantes (DGGE, por sus siglas en inglés). Además, se identificaron las poblaciones por secuenciación de las bandas dominantes en los perfiles de DGGE. La evolución de la biodegradación se estimó a través de la actividad deshidrogenasa durante los ensayos. El análisis molecular de los extractos de ADN, reveló una riqueza de genotipos similar en ambos tratamientos recién contaminados y una disminución importante de este parámetro por efecto de intemperismo. Los niveles máximos de actividad de la enzima se presentaron en el suelo recién contaminado-bioestimulado (3146.84 mg de iodo-nitro-formazan (INF)/g) en comparación con el suelo recién contaminado bajo atenuación natural (1160.19 µg INF/g). Para los suelos intemperizados, la actividad deshidrogenasa disminuyó en comparación con los demás tratamientos. Se determinó una comunidad bacteriana estable a lo largo del tiempo en el suelo intemperizado, mientras que la adición de nutrientes favoreció el incremento de la riqueza de genotipos tanto en los suelos recién contaminados como en los intemperizados. Se identificó la presencia de bacterias hidrocarbonoclastas (Burkholderia, Pelobacter) en todos los tiempos de muestreo en ambos tratamientos.Key words: enzymatic activity, DGGE, soil restoration, sequencing ABSTRACT Dehydrogenase activity and composition of the native microbial community were evaluated in hydrocarbon-contaminated soil during its bioremediation by biostimulation Rev. Int.

show abstract

A Simple Silica-based Method for Metagenomic DNA Extraction from Soil and Sediments

Cited by 51 publications

References 18 publications

New efficient DNA extraction method to access the microbiome of Ricinus communis seeds

New efficient DNA extraction method to access the microbiome of Ricinus communis seeds

Methodology Simple and inexpensive DNA extraction protocol for studying the bacterial composition of sludges used in microbial fuel cells

Dinámica De Poblaciones Bacterianas Y Actividad Deshidrogenasa Durante La Biorremediación De Suelo Recién Contaminado E Intemperizado Con Hidrocarburos

Contact Info

Product

Resources

About