We are using commercial kits but in some cases are an expensive approach for initial screenings. I need a buffer for was the sample, remove PCR inhibitors and sediment major particles. After this first wash maybe conventional kits could be used successfully.
Currently in our laboratory we have developed a system of isolate bacterial DNA from stool that combines a standard protocol of DNA isolation with phenol-chloroform and a second part in which we use commercial kits to remove all inhibitory substances to the sample. The truth is the only process we have found that we get the sufficient amounts of DNA with a very good efficiency (ratio 260/280 above 1.8). If you give me an email address I could send you a copy of our protocol.
Regards and good luck with the isolation because faeces sample are very difficult to work
Hi
Send the protocol for me too please ([email protected])
Regards
Diogo Pinho
Ok, I can provide the protocol but at the moment it is only in Spanish. If you do not understand something you can contact me.
Regards
Hello,
The following paper describes a very effective method for DNA extraction from the fecal samples. We are using this method and always get good DNA quality. Hope it works for you. Best regards.
Huyen Tran
Diet-Induced Metabolic Improvements in a Hamster Model of Hypercholesterolemia Are Strongly Linked to Alterations of the Gut Microbiota. Inés Martínez, Grant Wallace, Chaomei Zhang, Ryan Legge, Andrew K. Benson, Timothy P. Carr, Etsuko N. Moriyama, and Jens Walter. Appl. Environ. Microbiol. 2009, 75(12):4175. DOI: 10.1128/AEM.00380-09.
thanks a lot ,
Really all the procedures reviewed are complex and not easily automatize. Maybe if during the first washing steps we use specific resins we can reduce the work.
I try to work with these procedures.
Hi..on my practical and research experiences,Isolating pure and quality DNA you can opt Phenol -Chloroform protocol.
With warm regards..
thanks Seweryn !
then this lost log, is related with partial PCR inhibition ?
Hi, Antonio Galiana, could send me your protocol of isolate bacterial DNA from stool, please. We initiated work related to Enterobacterial in human stool samples. our interest is specifically detected by PCR at several genres such com Escherichia, Salmonella, Shigella. We developed some procedures to identify these pathogens.
We are in Mexico City and work in a diagnostic center and surveillance.
Best regards.
Try the new PureLink Microbiome DNA purification kit - it has protocols for soil, stool, etc. works really well for microbiome applications. not trivial to obtain DNA without good purification- as you need to deplete bile, bilirubin, humic acids etc. Additional PCR boost can be accomplished by addition of BSA in PCR reaction or by using "tough" enzymes- that can work in the presence of inhibitors
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