FIG. 3.

Mature double-stranded DNA synthesis and deproteinization occur in purified intracellular DHBV nucleocapsids following endogenous DNA polymerase reaction. (A) EPRs were performed with immature intracellular nucleocapsids prepared from Dstet5 cells (see Materials and Methods for details) in a 100-μl EPR mixture as described in the legend of Fig. 1. The reaction mixtures were either without incubation (lane 2) or incubated at 37°C for the indicated periods of time (lanes 3 to 6). Core DNA and DP DNA were analyzed by Southern blot hybridization. The amount of core DNA (upper panel) and DP DNA (lower panel) loaded onto each lane was derived from 4 × 10⁶ and 4 × 10⁷ cells. (B) EPRs were performed with immature intracellular DHBV nucleocapsids, and the reaction mixtures were either without incubation (lanes 2 and 6) or incubated at 37°C for the indicated periods of time (lanes 3 to 5 and 7 to 11). Core DNA and DP DNA were extracted without (lanes 2 to 9) or with (lanes 10 and 11) prior DNase I digestion and detected by Southern blot hybridization with a DHBV minus-strand specific α-³²P-riboprobe. Fifty picograms of 3.0-kb unit-length DHBV DNA was loaded as the hybridization size marker (lane 1). The positions of rcDNA (RC), dslDNA (DSL), single-stranded DNA (SS), and partial single-strand DNA are indicated.