Abstract: Based on the published sequences of picornavirus, 4 pairs of RT-PCR primer were designed. Total RNA were extracted from chick embryo allantoic fluid by RNAiso Reagent,one cDNA fragment were successfully amplified from total RNA extracted from DHV A₆₆chick embryo allantoic fluid by using TakaRa
RNA PCR Kit（AMV）Ver3.0. After ligating cDNA to the plasmid vector pMD18T and introducing into E. coli DH5а, the recombinant plasmids were screened by ampicillin and checked by the restriction enzyme EcoRⅠ and SalⅠ digestion. Then the recombinant plasmids were sequenced. Compare and analyse of homologies of nucleotide showed that identities of DHV A₆₆ and 3 published DHV were up to 96.2%～96.8%. This indicated partial cDNA library of DHV A₆₆ strain were constructed successfully.

Key words: duck hepatitis virus(DHV); A₆₆strain; cDNA library; sequence analysis