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Bacterial DNA extraction/purification from animal tissue - (May/25/2014 )



I would like to extract/purify *bacterial* DNA from animal tissues for bacterial identification through 16S PCR, including probably gram positive bacteria.


I have QIAGEN DNeasy blood/tissue kits. There are two protocols avaliable, one for tissues, and another one for gram positive bacteria.


I wonder which protocol should I follow, since I am not working with bacterial suspensions proper, and in the other hand I do not want to miss gram positives due to lysis failure.


I thought to add lysozime to the lysis step, but then I am not sure the chemistry, pH, temperature etc. are suitable for lysozime.


I found a paper suggesting an additional sonication step is done. But they did not test it with QIAGEN's kit, and they do not specify at which step to add sonication.


Any suggestions? Thanks a lot.


A more certain protocol would be to use phenol/chloroform extraction, perhaps preceded by a lysozyme or lysostaphin digestion. Phenol chloroform will likely get everything with DNA in it, while the kits may miss things.