[Construction of infectious molecular clone of duck hepatitis virus type 1 CL strain].

OBJECTIVE

To construct an infectious clone for studying functions of duck hepatitis virus (DHV) typel genome by reverse genetic technique.

METHODS

Three fidelity DNA fragments covering the full genome of DHV type 1 CL strain were amplified by RT-PCR, and inserted into pBR322 vector, resulting in the full-length cDNA clone BR-CL. The in vitro-transcribed RNA from BR-CL was transfected into duck embryo renal cells and the rescued virus was identified using RT-PCR, indirect immunofluorescence assay and colloidal gold immunoelectron microscopy after six generations. After inoculating the rescued virus into SPF chick embryos, embryo death and pathological changes were observed.

RESULTS

The results of RT-PCR, indirect immunofluorescence assay and immunoelectron microscope showed that infectious virus was rescued. After inoculating into SPF chick embryos, the rescued virus was able to kill embryos with pathogenic changes.

CONCLUSION

This is the first report on generation of infectious cDNA clone of DHV, which provides a valuable platform for further research on functions of DHV genome.