Beet soil-borne virus RNA 1: genetic analysis enabled by a starting sequence generated with primers to highly conserved helicase-encoding domains.

Abstract

The complete sequence of the 5834 nucleotides of RNA 1 of beet soil-borne furovirus (BSBV, Ahlum isolate) was determined using a PCR product obtained with primers to highly conserved coding regions for helicase-like proteins in RNA 1 of furo-, hordei-and tobraviruses as a starting sequence. Unknown parts of the sequence upstream and downstream of this starting sequence were amplified by means of RT-PCR techniques using combinations of specific and random primers. BSBV RNA 1 contains one large ORF for a readthrough protein with a molecular mass of 204 kDa (204K protein) which is interrupted internally by a UAA stop codon terminating the coding region for a protein of 145 kDa (145K protein). The N-and C-terminal parts of the 145K protein and the readthrough domain of the 204K protein contain methyltransferase, helicase and RNA-dependent RNA polymerase motifs, respectively. Unlike other furoand tobraviruses BSBV contains no further genes on its RNA 1.

Genome properties of beet virus Q, a new furo-like virus from sugarbeet, determined from unpurified virus.

“…Because BVQ is extremely difficult to purify, a strategy was adopted for analysing its genome properties which is based partially on previous experiences with BSBV (Koenig, 1997 ;Koenig & Loss, 1997 ;Koenig et al, 1997) and is outlined in Fig. 1.…”

“…A starter sequence, which was generated with primers specific for highly conserved helicase-encoding domains in RNA 1 of furo-, pomo-, peclu-, hordei-and tobraviruses (Koenig & Loss, 1997), has enabled the entire nucleotide sequence of the tripartite genome of unpurified BVQ to be determined. This paper describes genome properties of BVQ and compares them with those of other, related viruses.…”

A full-length infectious clone of beet soil-borne virus indicates the dispensability of the RNA-2 for virus survival in planta and symptom expression on Chenopodium quinoa leaves

“…Originally reported in Italy (Canova, 1959), rhizomania disease is now widespread in most countries where sugar beet is grown (McGrann et al, 2009) and BSBV is often found in beet infected with BNYVV (Meunier et al, 2003). However, the pathogenicity of BSBV and its contribution to the rhizomania syndrome remain unclear, with opinions still divided on this (Prillwitz & Schlösser, 1992;Kaufmann et al, 1993;Lindsten, 1993;Rush & Heidel, 1995).The BSBV genome consists of three single-stranded RNAs of positive polarity, packaged into rod-shaped particles (Koenig et al, 1996(Koenig et al, , 1997Koenig & Loss, 1997). RNA-1 (5.8 kb) encodes the putative viral replicase.…”

“…The BSBV genome consists of three single-stranded RNAs of positive polarity, packaged into rod-shaped particles (Koenig et al, 1996(Koenig et al, , 1997Koenig & Loss, 1997). RNA-1 (5.8 kb) encodes the putative viral replicase.…”

“…After total RNA extraction from sugar beet roots using the SV total RNA isolation kit (Promega), the three genomic RNAs were reverse transcribed and amplified by PCR (RT-PCR) using expand reverse transcriptase and the expand long template PCR System (Roche). Primers matching the extremities of the three viral RNAs were designed following the only references known to date (Koenig et al, 1996(Koenig et al, , 1997Koenig & Loss, 1997). Forward primers were flanked with a T7 promoter sequence (TAATACGACTCACTATAG) (Fig.…”