Efficient protocols for the extraction of microbial DNA from the rhizosphere of hydrophilic forests in Chile

A lysis buffer-based protocol (Protocol BA), a modified lysis buffer-based protocol (Protocol BA Mod) and a commercial extraction kit (Protocol PS Kit) (Power Soil, Mo bio Laboratories, CA USA) were each evaluated for their ability to produce high-quality DNA with yields sufficient to allow its use...

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Autores principales:	Cáceres,Pablo, Cordero,Cecilia, González,Gloria, Quiroz,Karla, Bobadilla,Juan C, Bravo,Carmen, Caligari,Peter D.S, Carrasco,Basilio, García-Gonzales,Rolando
Lenguaje:	English
Publicado:	Pontificia Universidad Católica de Chile. Facultad de Agronomía e Ingeniería Forestal 2012
Materias:	hydrophilic forests 16S ribosomal RNA PCR rhizosphere soil DNA extraction
Acceso en línea:	http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0718-16202012000300018
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spelling	oai:scielo:S0718-162020120003000182014-03-10Efficient protocols for the extraction of microbial DNA from the rhizosphere of hydrophilic forests in ChileCáceres,PabloCordero,CeciliaGonzález,GloriaQuiroz,KarlaBobadilla,Juan CBravo,CarmenCaligari,Peter D.SCarrasco,BasilioGarcía-Gonzales,Rolando hydrophilic forests 16S ribosomal RNA PCR rhizosphere soil DNA extraction A lysis buffer-based protocol (Protocol BA), a modified lysis buffer-based protocol (Protocol BA Mod) and a commercial extraction kit (Protocol PS Kit) (Power Soil, Mo bio Laboratories, CA USA) were each evaluated for their ability to produce high-quality DNA with yields sufficient to allow its use in biodiversity studies. Similarly, the effect of liquid nitrogen on the process of cell disruption in all of the protocols that were studied. DNA yields ranged from 12.4 ng g-1 of processed soil to 9620 ng g-1 using the modified lysis buffer and commercial extraction kit, respectively. The quality of the DNA was determined by the ability of the DNA to produce efficient and reproducible polymerase chain reaction (PCR) products, using primers for universal 16S and 18S ribosomal RNA regions from bacteria and fungi, respectively. High-quality DNA was obtained to run PCRs in all protocols, but the efficiency of the method depended on the dilution of the DNA prior to performing the PCR. The three extraction methods generated PCR products with 90% efficiency. The DNA produced with the commercial kit was able to produce the highest PCR efficiency (95%) when the 10-1 dilution was used. The method based on the use of lysis buffer produced the highest efficiency (90%) using a 10-2 dilution. Meanwhile, the modified lysis buffer-based protocol generated the highest efficiency of PCR products using the 10-3 dilution factor with 95% of efficiency. For the first time, reliable and efficient DNA isolation from the rhizosphere of hydrolic forest is documented, enabling a wide range of applications for this technique.info:eu-repo/semantics/openAccessPontificia Universidad Católica de Chile. Facultad de Agronomía e Ingeniería ForestalCiencia e investigación agraria v.39 n.3 20122012-12-01text/htmlhttp://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0718-16202012000300018en10.4067/S0718-16202012000300018
institution	Scielo Chile
collection	Scielo Chile
language	English
topic	hydrophilic forests 16S ribosomal RNA PCR rhizosphere soil DNA extraction
spellingShingle	hydrophilic forests 16S ribosomal RNA PCR rhizosphere soil DNA extraction Cáceres,Pablo Cordero,Cecilia González,Gloria Quiroz,Karla Bobadilla,Juan C Bravo,Carmen Caligari,Peter D.S Carrasco,Basilio García-Gonzales,Rolando Efficient protocols for the extraction of microbial DNA from the rhizosphere of hydrophilic forests in Chile
description	A lysis buffer-based protocol (Protocol BA), a modified lysis buffer-based protocol (Protocol BA Mod) and a commercial extraction kit (Protocol PS Kit) (Power Soil, Mo bio Laboratories, CA USA) were each evaluated for their ability to produce high-quality DNA with yields sufficient to allow its use in biodiversity studies. Similarly, the effect of liquid nitrogen on the process of cell disruption in all of the protocols that were studied. DNA yields ranged from 12.4 ng g-1 of processed soil to 9620 ng g-1 using the modified lysis buffer and commercial extraction kit, respectively. The quality of the DNA was determined by the ability of the DNA to produce efficient and reproducible polymerase chain reaction (PCR) products, using primers for universal 16S and 18S ribosomal RNA regions from bacteria and fungi, respectively. High-quality DNA was obtained to run PCRs in all protocols, but the efficiency of the method depended on the dilution of the DNA prior to performing the PCR. The three extraction methods generated PCR products with 90% efficiency. The DNA produced with the commercial kit was able to produce the highest PCR efficiency (95%) when the 10-1 dilution was used. The method based on the use of lysis buffer produced the highest efficiency (90%) using a 10-2 dilution. Meanwhile, the modified lysis buffer-based protocol generated the highest efficiency of PCR products using the 10-3 dilution factor with 95% of efficiency. For the first time, reliable and efficient DNA isolation from the rhizosphere of hydrolic forest is documented, enabling a wide range of applications for this technique.
author	Cáceres,Pablo Cordero,Cecilia González,Gloria Quiroz,Karla Bobadilla,Juan C Bravo,Carmen Caligari,Peter D.S Carrasco,Basilio García-Gonzales,Rolando
author_facet	Cáceres,Pablo Cordero,Cecilia González,Gloria Quiroz,Karla Bobadilla,Juan C Bravo,Carmen Caligari,Peter D.S Carrasco,Basilio García-Gonzales,Rolando
author_sort	Cáceres,Pablo
title	Efficient protocols for the extraction of microbial DNA from the rhizosphere of hydrophilic forests in Chile
title_short	Efficient protocols for the extraction of microbial DNA from the rhizosphere of hydrophilic forests in Chile
title_full	Efficient protocols for the extraction of microbial DNA from the rhizosphere of hydrophilic forests in Chile
title_fullStr	Efficient protocols for the extraction of microbial DNA from the rhizosphere of hydrophilic forests in Chile
title_full_unstemmed	Efficient protocols for the extraction of microbial DNA from the rhizosphere of hydrophilic forests in Chile
title_sort	efficient protocols for the extraction of microbial dna from the rhizosphere of hydrophilic forests in chile
publisher	Pontificia Universidad Católica de Chile. Facultad de Agronomía e Ingeniería Forestal
publishDate	2012
url	http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0718-16202012000300018
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Efficient protocols for the extraction of microbial DNA from the rhizosphere of hydrophilic forests in Chile

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