[Cloning and phylogenetic analysis of hepatitis-associated TT virus subgenome]

Polymerase chain reaction (PCR) was utilized for the DNA amplification from transfusion transmitted virus (TTV) positive serum samples. Five TTV DNA fragments, overlapped about 90% of the genome, were amplified by long template PCR for the generation of TTV subgenome. Recombinant plasmids were obtained by directly inserting PCR products into pT-Adv vector, and DNA sequence analyses showed they were TTV DNA fragments. By using specific restriction enzymes, five TTV DNA fragments were ligated into a TTV DNA subgenome clone and named as TTV021. TTV021 has been deposited in GenBank database with the accession number AF254410. The results of computer analyses showed that TTV021, 3472 nt long, contains two open reading frames (ORF1, 785 aa; ORF2, 146 aa). Identity alignments between TTV021 and other TTV isolates indicated several high conserved regions existed. Phylogenetic analysis of 356 nt from TTV021 suggested that the isolate has close evolutionary relationship with CHN1 (type 1a), but has far relation with other TTV isolates.