A simple method of DNA extraction for Eimeria species

A new, simple method is described for extracting DNA from coccidia (Eimerii dae) oocysts. In our hands this method works well for all Eimeria oocysts a nd, presumably, will work equally well for oocysts of other coccidia genera . This method combines the two steps of breaking oocyst and sporocyst walls , and dissolving the sporozoite membrane in one step. This greatly simplifi es the currently used DNA extraction procedures for Eimeria species and ove rcomes the disadvantages of existing DNA extraction methods based on glass- bead grinding and sporozoite excystation procedures. Because all the proced ures are done in a 1.5-ml microfuge tube. which minimizes the loss of DNA i n the extraction procedures, this method is especially suitable for samples with small number of oocysts. In addition, this method directly lyses the oocyst and sporocyst walls as well as the sporozoite membrane in a continuo us incubation; therefore, it does not require the sporozoites to be alive. The results of PCR experiments indicate that this method generates better q uality of DNA than what the existing glass-bead grinding method does for mo lecular analysis, and is suitable for both large or small number(< 10(2) oo cysts) of living or dead oocyst samples. (C) 2001 Published by Elsevier Sci ence B.V.