FIG. 4.

Fractionation of the cytoplasmic RNP complexes isolated from plants infected with TMV(ORF3) by Cs₂SO₄ density gradient centrifugation. (A) Diagram showing density (ρ) gradient in the fractions obtained after centrifugation. The positions of the rods and disks (rods) and cytoplasmic RNP complexes (RNP-C) determined by electron microscopy and of fractions possessing infectivity are indicated. (B) Western blot analysis of the fractions, using antibody against the GRV ORF3 protein. The positions of the dimers (d) and monomers (m) of the ORF3 protein are indicated. Fraction b (bottom) is a sample of the resuspended pellet from the gradient. (C) Electrophoretic protein analysis (SDS-PAGE) of a partially purified preparation of the cytoplasmic RNP complexes (pooled fractions 11 to 14). Gel was stained with Coomassie blue (lane 1) or analyzed by Western blotting as in panel B (lane 2). The positions of the molecular mass markers (M) and of the dimers (d) and monomers (m) of the ORF3 protein are indicated to the right and left of the gel, respectively. (D) Dot blot hybridization analysis of TMV(ORF3) RNA contained in combined fractions 3 to 6, 7 to 10, and 11 to 14. Hybridization was with a probe specific to the ORF3 nucleotide sequence.