Data/hora: 09/06/2024 09:35:42

Provedor de dados: 	Institutional Archive of Ifreme (ArchiMer)	
País: 	France	
Título: 	DNA extractions from deep subseafloor sediments: Novel cryogenic-mill-based procedure and comparison to existing protocols	
Autores: 	Alain, Karine; Callac, Nolwenn; Ciobanu, Maria Cristina; Reynaud, Yann; Duthoit, Frederique; Jebbar, Mohamed.	
Data: 	2011-12	
Ano: 	2011	
Palavras-chave: 	Sediment; DNA extraction; Deep subsurface biosphere.	
Resumo: 	Extracting DNA from deep subsurface sediments is challenging given the complexity of sediments types, low bio-masses, resting structures (spores, cysts) frequently encountered in deep sediments, and the potential presence of enzymatic inhibitors. Promising results for cell lysis efficiency were recently obtained by use of a cryogenic mill (Lipp et al., 2008). These findings encouraged us to devise a DNA extraction protocol using this tool. Thirteen procedures involving a combination of grinding in liquid nitrogen (for various durations and beating rates) with different chemical solutions (phenol, chloroform, SDS, sarkosyl, proteinase, GTC), or with use of DNA recovery kits (MagExtractor (R)) were compared. Effective DNA extraction was evaluated in terms of cell lysis efficiency, DNA extraction efficiency, DNA yield and determination of prokaryotic diversity. Results were compared to those obtained by standard protocols: the FastDNA (R) SPIN kit for soil and the Zhou protocol. For most sediment types grinding in a cryogenic mill at a low beating rate in combination with direct phenol-chloroform extraction resulted in much higher DNA yields than those obtained using classical procedures. In general (except for clay-rich sediments), this procedure provided high-quality crude extracts for direct downstream nested-PCR, from cell numbers as low as 1.1 x 10(6) cells/cm(3). This procedure is simple, rapid, low-cost, and could be used with minor modifications for large-scale DNA extractions for a variety of experimental goals. (C) 2011 Elsevier B.V. All rights reserved.	
Tipo: 	Text	
Idioma: 	Inglês	
Identificador: 	http://archimer.ifremer.fr/doc/00056/16691/14364.pdf

DOI:10.1016/j.mimet.2011.09.015	
Editor: 	Elsevier Science Bv	
Relação: 	http://archimer.ifremer.fr/doc/00056/16691/	
Formato: 	application/pdf	
Fonte: 	Journal Of Microbiological Methods (0167-7012) (Elsevier Science Bv), 2011-12 , Vol. 87 , N. 3 , P. 355-362	
Fonte: 	2011 Elsevier B.V. All rights reserved.