Abstract

Total RNA of Eimeria tenella drug-resistant strain from Tangshan was extracted with Trizol. Differential-display reverse transcription- polymerase chain reaction (DDRT-PCR) was established by 3 anchored primers and 20 arbitrary primers. The products ofpolymerase chain reaction (PCR) were analyzed on the denaturing polyacrylamide gels by silver-straining. Ten differential bands were excised from the gels and reampilfied with the same sets of primers. The products were purified and ligated with PMD^TM18-T Vector, and then the dot-blot hybridization, sequence analysis and homology comparison. The results showed that through comparison of the nucleotide acid sequence, the similarity was 99% among the sequence S116 from mRNA of Tangshan multiple-resistant strain with the sequence 882 bp lengths in the first chromosome of E.tenella in Genebank and Sanger, which was an unknown protein. This study paved the way for cloning the full-length cDNAs (Complementary Deoxyribonucleic acid) and finding the molecular mechanism about the drug-resistance of E.tenella.

Key words: Chicken, E. tenella, mRNA differential display PCR, denaturing polyacrylamide gel, gene cloning.