To increase DNA output from microsporidia (usually presented by multiple spores with rigid spore wall) from host tissues (dry cadavers or fresh bodies of pyraloid moths) I usually homogenize sample in distilled water and filter it through a piece of cotton (in a syringe) prior to DNA extraction. I believe that partial removal of host tissues increases presence of parasite's spores in sample and subsequent DNA output, at least the specific PCR signal after such procedure is increased. This time, however, the sample is in 1% SDS Tris buffered solution. Can I treat it as if it was in water, just homogenize and filter, then extract DNA? Or partial tissue lysis in these samples (including the parasite spores) will prevent good results?
I think if you treat this 1% SDS Tris buffer as water it seems ok and it will work in PCR.
Dear....,
you need not filter the homogenate to limit host tissue....Even in your normal method, you need not go for this filtration. Because, PCR by design and theory amplifies only the specific and targeted DNA irrespective of the presence of other DNA populations unless you are using degenerate primers or heavy concentration of DNA.
All the best for your research....
@Binech C.P,
I am more than sure many will argue that "PCR .... amplifies only the specific and targeted DNA irrespective of the presence of other DNA ". Be that truth, there would be no need to design protocols like touch down PCR - another approach that I hopefully gonna use to solve my problem
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