RNA extraction and Northern Blot
The total RNAs for northern blot were extracted using hot phenol method (Kohrer and Domdey, 1991). Then equal amount (15 μg) of RNAs were used for electrophoresis and transferred to nylon membranes. Digoxigenin (DIG)-labeled probes targeting the CP-coding sequence of ApNMV were generated and used for hybridization to detect the positive-strand RNA3 ((+)RNA3) and (+)RNA4 per manufacture’s protocol (Roche). Anti-DIG antibody (conjugated with alkaline phosphatase, Roche) was used for immunoblotting and a chemiluminescent substrate for alkaline phosphatase was applied for imaging through a gel imager (Bio-RAD).