TABLE 1.
Properties of particles and RNA found in CNV CP mutants
| Mutanta | %WT CNV yieldb | T=3:IS:T=1 ratioc | Mass of CP (kDa)d
|
Size (kb) of RNA species found in particles of CNV mutantse
|
||
|---|---|---|---|---|---|---|
| Predicted | Observed | Predicted | Observed | |||
| WT CNV | 100 | 1:0:0 | 41 | 41 | 4.7 (2.09) | 4.7 |
| P85G | 0.4 | 1:0:1.5 | 41 | 41, 34, 31 | 4.7 | 4.7, 0.7 |
| R1+2 | 2.8 | 1:0:1.5 | 36 | 36, 34 | 4.58 (1.97) | 1.9, 1.4, 0.9 |
| R1+2/P85G | 5.8 | 1:0:10 | 36 | 36, 34 | 4.58 | 0.6-0.75 |
| R3(−) | 14.9 | 7:7:1 | 40 | 40, 38, 34, 31 | 4.68 (2.08) | 2.1, 1.2-1.5, 0.6-0.75 |
| R3(−)/p85G | 0.7 | 0:1:1.4 | 40 | 40, 34, 31 | 4.68 | 0.6 |
| RΔ32-58 | 6.5 | >95% T=1 | 38 | 38, 37, 34 | 4.62 (2.02) | 2.0, 0.6 |
| RΔ2-58 | 0.6 | >95% T=1 | 34 | 34 | 4.53 | 0.6-0.75 |
See Fig. 2 for description of mutants.
Percent WT CNV yield indicates the yield of mutant particles as a percentage of that of WT CNV. Equivalent masses of infected leaves were used, and the values shown are the average yields in mass obtained from two independent experiments.
The approximate T=3:IS:T=1 particle ratio was assessed by TEM of uranyl acetate-stained particles. At least 250 particles were examined using several different areas of one or more TEM grids. In the case of P85G, particle ratios were obtained only from preparations that contained T=1 particles (see text).
Predicted masses are based on the known mass of the CNV CP (40) minus the mass of the amino acids deleted in each construct. Observed masses were estimated following denaturing polyacrylamide gel electrophoresis as described for Fig. 6 and 8. Numbers in italics correspond to the approximate sizes of deleted CP species found in the various mutants. Underlined numbers correspond to CP species that are enriched in IS or T=1 particles of the indicated mutant (Fig. 8). The 38-kDa species was observed only for IS particles of R3(−).
The known sizes of WT CNV gRNA (40) and the predicted sizes of the gRNA of each of the mutants are shown in kilobases. For mutants observed to encapsidate an RNA species similar in size to CNV sgRNA1 (2.09 kb) (34), the predicted size of the sgRNA1 for that mutant is shown in parenthesis. Observed sizes correspond to the approximate sizes of the predominant RNA species extracted from particle preparations. Species that may correspond to encapsidated sgRNA1 are shown in italics. Sizes were estimated following denaturing agarose gel electrophoresis as described in Fig. 7. The 0.7 kb RNA of P85G was not always present in particle preparations (see text).