Subcellular localization of tobacco mosaic virus minus strand RNA in infected protoplasts.
Radioactive RNA probes were prepared which specifically hybridize with sequences complementary to 5' and 3' regions of tobacco mosaic virus (TMV) RNA. These probes were used in Northern hybridization to locate TMV-RNA minus strands in the subcellular fractions of infected tobacco protoplasts. When the protoplasts were lysed with Triton X-100, full-length minus strands were present in the cytoplasmic but not in the nuclear fraction. With mechanically broken protoplasts, the crude nuclear fraction (250 g pellet) contained small amount of minus strands which appeared to derive from unbroken protoplasts, but most of minus strands were recovered in a fraction sedimented between 250 and 2500 g, little if any being found in lighter fractions. The results indicate that TMV-RNA replicates in association with an extranuclear structure.
Okamoto
S
S
Department of Biology, Faculty of Science, Nagoya University, Japan.
Machida
Y
Y
Takebe
I
I
D015152:Blotting, Northern; D002458:Cell Fractionation; D002460:Cell Line; D002467:Cell Nucleus / Q000032:analysis; D002499:Centrifugation, Density Gradient; D003593:Cytoplasm / Q000032:analysis; D009693:Nucleic Acid Hybridization; D010947:Plants, Toxic; D011523:Protoplasts / Q000032:analysis*; D015347:RNA Probes; D012330:RNA, Double-Stranded / Q000032:analysis; D012367:RNA, Viral / Q000032:analysis* / Q000096:biosynthesis; D014026:Nicotiana; D014027:Tobacco Mosaic Virus / Q000235:genetics* / Q000648:ultrastructure
D015347:RNA Probes; D012330:RNA, Double-Stranded; D012367:RNA, Viral