Cloning of large positive-strand RNA viruses.
Full-length, biologically active cDNA clones of the positive-strand RNA plant viruses are indispensable for investigating the functions of viral genes and control elements as well as generating virus-derived gene expression and silencing vectors. Even though engineering of such clones for 4- to 10-kb viral RNAs has become routine, it remains a challenging task for 15- to 20-kb RNA genomes of the monopartite viruses in a family Closteroviridae. This unit describes strategic considerations and techniques used to generate an infectious cDNA clone of a closterovirus. The use of agroinfection to improve specific infectivity of the resulting clone is also explained.
V Peremyslov
Valera
V
Oregon State University, Corvallis, Oregon, USA.
V Dolja
Valerian
V
D016960:Agrobacterium tumefaciens / Q000235:genetics*; D003001:Cloning, Molecular / Q000379:methods*; D017790:Closterovirus / Q000235:genetics*; D018076:DNA, Complementary / Q000235:genetics; D005818:Genetic Engineering / Q000379:methods*; D012328:RNA Viruses / Q000235:genetics; D012367:RNA, Viral / Q000235:genetics* / Q000302:isolation & purification; D014026:Nicotiana / Q000821:virology*; D014169:Transformation, Bacterial
D018076:DNA, Complementary; D012367:RNA, Viral