The complete nucleotide sequence of an infectious clone of soybean chlorotic mottle virus (SoyCMV) DNA was determined and compared with those of three other caulimoviruses, cauliflower mosaic virus (CaMV), carnation etched ring virus and figwort mosaic virus. The double-stranded DNA genome of SoyCMV (8,175 bp) contained nine open reading frames (ORFs) and one large intergenic region. The primer binding sites, gene organization and size of ORFs were similar to those of the other caulimoviruses, except for ORF I, which was split into ORF Ia and Ib. The amino acid sequences deduced from each ORF showed only short, highly homologous regions in several of the corresponding ORFs of the three other caulimoviruses. A promoter fragment of 378 bp in SoyCMV ORF III showed a strong expression activity, comparable to that of the CaMV 35S promoter, in tobacco mesophyll protoplasts as determined by a beta-glucuronidase assay using electrotransfection. The fragment contained CAAT and TATA boxes but no transcriptional enhancer signal as reported for the CaMV 35S promoter. Instead, it had sequences homologous to a part of the translational enhancer signal reported for the 5'-leader sequence of tobacco mosaic virus RNA.
A tospovirus-like virus recovered from netted melon was transmitted by Thrips palmi in a persistent manner but had different cytopathological features from tospoviruses previously reported. Viral nucleocapsid (N) was purified with two protective reagents, 2-mercaptoethanol and L-ascorbic acid, and RNA extracted from the viral nucleocapsid was used for genomic analysis. The virus had a genome consisting of three single-stranded RNA molecules. The open reading frame on the viral complementary strand, located at the 3' end of the viral S RNA, encoded the N protein. The 3' terminus of this RNA also contained an eight-nucleotide sequence similar to the conserved sequence at the 3' end of genomic RNA molecules of tospoviruses. These features of the viral genome are identical to those of tospoviruses; therefore, this virus is considered to belong to the genus Tospovirus. Its N protein comprised 279 amino acids and had a molecular mass of 31.0 kDa. Comparisons of its amino acid sequence with those of known tospoviruses revealed less than 60% identity. This melon virus is concluded to be a distinct species in the genus Tospovirus, and the name Melon yellow spot virus is proposed.
A semiselective medium (PCCG) and a polymerase chain reaction (PCR) technique were combined to detect viable cells of Ralstonia (Pseudomonas) solanacearum E. F. Smith (synonym Burkholderia solanacearum) in soil. DNA was extracted from 92 strains of soil bacteria including R. solanacearum that grew on PCCG and then used as template for PCR with a pair of primers designed to amplify a single fragment (281 bp) of R. solanacearum DNA. The 281‐bp fragment was amplified only from DNA of R. solanacearum (12 strains). For DNA from soil bacteria other than R. solanacearum, the PCR amplification generated no products for 66 strains, a single DNA band with different sizes from 281 bp for 2 strains, and several DNA bands for 12 strains. Southern analysis showed that any of those products other than the 281‐bp fragment had no homology with the 281 ‐bp fragment of R. solanacearum, indicating the specificity of the primers to generate the 281‐bp fragment from R. solanacearum. A simple method consisting of a plating step using PCCG and succeeding PCR for amplifying the 281‐bp fragment from colonies on PCCG was described.
The complete nucleotide sequence of RNA-2 from a Japanese isolate IP of broad bean wilt virus (BBWV) was determined. The sequence encodes a single large polyprotein, which contains a putative movement protein and two coat proteins (CPs). The 3'-terminal sequences of RNA-2 were also determined for three other Japanese isolates and two ATCC isolates (PV132 and PV176) of BBWV. The CPs of the four Japanese isolates share 86.8-98.0% amino acid sequences homology with one another and 88.3-96.5% with those reported for the isolate PV131 (BBWV-2). However, they have only 57.9-66.2% homology with those of PV132 and PV176 (BBWV-1).
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