Construction of Subtractive cDNA Libraries of the Sporogony Stage of Eimeria tenella by Suppression Subtractive Hybridization

To clone and identify differentially expressed genes in the sporogony stage of Eimeria tenella, the cDNAs from unsporulated oocysts and sporulated oocysts of E. tenella were used as the driver, respectively, the cDNAs from sporozoites of E. tenella was used the tester. Two subtractive cDNA libraries of sporozoites were constructed using the suppression subtractive hybridization (SSH) technique. The cDNAs from unsporulated oocysts was used the driver, the cDNAs from sporulated oocysts was used the tester and one subtractive cDNA library of sporulated oocysts was constructed. PCR amplification revealed that the two subtractive cDNA libraries of sporozoites and one subtractive cDNA library of sporulated oocysts contained approximately 96%, 96% and 98% recombinant clones, respectively. Fifty positive clones were sequenced and analyzed in GenBank with Blast search from three subtractive cDNA libraries, respectively. Thirteen unique sequences were found from the subtractive cDNA library of sporulated oocysts, eight ESTs shared significant identity with the former. A total of 40 unique sequences were obtained from the two subtractive cDNA libraries, nine ESTs shared significant identity with the former and the other sequences represent novel genes of E. tenella with no significant homology to the proteins in Genbank. These results have provided the foundation for cloning new genes of E. tenella and further studying new approaches to control coccidiosis.