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Open Access item Lyophyllization improves the extraction of PCR-quality community DNA from pig faecal samples
Rubio, Luis A.
|Keywords:||lyophilisation, DNA, faecal microbiota, PCR–RFLP|
|Abstract:||BACKGROUND. Faeces are increasingly used as sources of DNA for genetic and ecological studies. Although multiple methods to preserve faecal samples prior to DNA extraction have been used (e.g. 70 % or absolute ethanol, freezing at -20ºC or in liquid nitrogen) no information is at present available in the literature on the use of lyophilized faeces. Accordingly, the yield and quality of the community DNA obtained by using four different commercial DNA extraction kits (QIAamp DNA Stool Mini Kit, REALPURE Spin Kit, SPEEDTOOLS Tissue DNA Extraction Kit, and JETQUICK Tissue DNA Spin Kit) from fresh and lyophilized samples of faeces were here studied. RESULTS. The use of lyophilized faeces resulted in a 1.5 to 2-fold increase in DNA recovery relative to the use of fresh faeces regardless of the kit used. Among the four kits tested, the best results were obtained with the QIAamp DNA Stool Mini Kit. Community DNA obtained from lyophilized faeces also provided the best restriction fragment length polymorphism (PCR-RFLP) profiles, which should guarantee a better representation of the microbial diversity present in faecal samples. CONCLUSION. As compared with using fresh faecal samples for pig faecal microbiota studies, lyophilization improved both DNA yield and quality of the information arising from the PCR- RFLP method of analysis.|
|Appears in Collections:||(EEZ) Artículos|
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