A plasmid-based transfection system capable of yielding infectious Porcine circovirus type 1 (PCV1) was established and mutational analysis was conducted to investigate the involvement of each viral transcription unit in protein synthesis, DNA replication and progeny virus production. During PCV1 replication in PK15 cells, twelve viral-specific RNAs are synthesized. They include the capsid protein RNA ( CR), eight Rep-associated RNAs ( Rep, Rep', Rep3a, Rep3b, Rep3c-1, Rep3c-2, Rep3c-3 and Rep3c-4), and three NS-associated RNAs ( NS462, NS642 and NS0). A stop codon introduced at the 5'-end of CR did not affect Rep-associated antigens or viral DNA synthesis. Altering the consensus dinucleotide at the splice junctions of the Rep3 RNAs and NS462 or introducing an early termination codon in Rep3c-4 and NS0 also did not have any affect on virus replication. However, mutations in Rep and Rep' caused greater than 99% reduction of protein synthesis and complete shut down of viral DNA replication. NS642 could not be assayed in this study because silent mutation at the splice junction was not possible. However, it is probably equivalent to the non-essential RNA ( NS672) of PCV type 2. Thus, only two proteins, Rep and Rep', are essential for PCV1 protein, DNA and infectious virus biosynthesis.