Molecular cloning and sequencing of the coat protein gene of a Nebraskan isolate of tobacco necrosis virus: the deduced coat protein sequence has only moderate homology with those of strain A and strain D

Abstract

A partial nucleotide sequence spanning the coat protein (CP) gene of a Nebraskan isolate of tobacco necrosis virus (TNV-NE) has been determined. The sequence contains at least four open reading frames (ORFs). The 5'-terminal ORF encodes a protein that has 86% and 38% homology with the polymerases of strains A (TNV-A) and D (TNV-D), respectively. The second and third ORFs probably encode 10.7 kDa and 6.2 kDa proteins (p 10.7 and p 6.2). These are respectively 90% and 96% amino acid homologous encoded by similar ORFs in TNV-A but only 26% and 20% homologous with those in TNV-D. The fourth 3'-proximal ORF encodes the 30.3 kDa CP. The amino acid sequence of TNV-NE CP is only 51% and 44% homologous to those of TNV-A and TNV-D, respectively. Thus, the CP genes of TNV-NE, TNV-A, and TNV-D are quite different. Like the sequences to the 5' side of the CP gene, that of TNV-NE is more closely related to TNV-A than to TNV-D.

Complete nucleotide sequence of tobacco necrosis virus strain DH and genes required for RNA replication and virus movement.

“…However, in the recently corrected TNV-D sequence a 7 kDa gene similar to the p7 " ORF of TNV-D H can be identified, though it was not mentioned by the authors . The complete genomic sequence of TNV-D H also allowed us to compare the amino acid sequences of each viral protein with all available sequences of TNV strains (A, D, D H ), the CP sequence of TNV-Nebraska (TNV-NE, Zhang et al, 1993) and two newly sequenced necroviruses : olive latent virus 1 (OLV-1 ; Grieco et al, 1996) and leek white stripe virus (LWSV ; Lot et al, 1996) and TCV (Table 1). From these comparisons, it is clear that TNV-D H has a high degree of sequence similarity with the other necroviruses, in particular with TNV-D.…”

“…TNV-D H -specific recombinant clones were sequenced by the dideoxy chaintermination method (Sanger et al, 1977) with T7 DNA polymerase (Sequenase, Amersham-US Biochemicals) on both strands of several independent cDNA clones. The sequence of the 5h region of TNV-D H RNA was determined by dideoxy- , TNV-NE (p6 and p11 ; Zhang et al, 1993), TNV-A (p6, p7 and p8 ; Meulewaeter et al, 1990), OLV-1 (p6 and p8 ; Grieco et al, 1996) and LSWV (p6 and p11 ; Lot et al, 1996)] and TCV carmovirus (p8 and p9 ; Carrington et al, 1989).…”

Evidence of olive mild mosaic virus transmission by Olpidium brassicae

“…The leaves were ground in cold 0.1 M sodium phosphate buffer (1:3 w/v), filtered, clarified in the presence of organic solvents, concentrated by differential centrifugation and further purified by ultracentrifugation through sucrose density gradient columns (Zhang et al 1993). The single light scattering virus band was recovered and concentrated by ultracentrifugation.…”

An efficient wheat transformation procedure: transformed calli with long-term morphogenic potential for plant regeneration

“…The resulting plasmid was cleaved with BamHI to remove the gus (uid A) gene from the plasmid, and the 5.2-kb product was religated followed by insertion of the nos terminator along with multiple cloning sites (MCS) from pBlueScript KSc. Wheat streak mosaic virus coat protein gene (cp), a mutant bacterial RNase III gene (rnc70 from plasmid pAM1551; Langenberg et al 1997), and tobacco etch virus P3 gene (P3) were amplified by PCR with primers including restriction sites, according to the procedure described by Zhang et al (1993). The annealing temperatures for PCR amplification of the selection marker gene and the three genes of interest were 63 7C (bar), 61 7C (cp), 70 7C (rnc70), and 60 7C (P3).…”