HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) Supplementary figures All Versions of this Article: vir.0.014712-0v1 91/2/552    most recent Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via CrossRef Citing Articles via Google Scholar Google Scholar Articles by Draghici, H.-K. Articles by Varrelmann, M. Search for Related Content PubMed PubMed Citation Articles by Draghici, H.-K. Articles by Varrelmann, M. Agricola Articles by Draghici, H.-K. Articles by Varrelmann, M.

Evidence for similarity-assisted recombination and predicted stem–loop structure determinant in potato virus X RNA recombination

Heidrun-Katharina Draghici^1,

Mark Varrelmann^1,2,

¹ Department of Crop Sciences, Section Plant Virology, University of Göttingen, Grisebachstrasse 6, D-37077 Göttingen, Germany
² Institute of Sugar Beet Research, Department of Phytopathology, Holtenser Landstrasse 77, D-37079 Göttingen, Germany

Correspondence
Mark Varrelmann
varrelmann{at}ifz-goettingen.de

Virus RNA recombination, one of the main factors for genetic variability and evolution, is thought to be based on different mechanisms. Here, the recently described in vivo potato virus X (PVX) recombination assay [Draghici, H.-K. & Varrelmann, M. (2009). J Virol 83, 7761–7769] was applied to characterize structural parameters of recombination. The assay uses an Agrobacterium-mediated expression system incorporating a PVX green fluorescent protein (GFP)-labelled full-length clone. The clone contains a partial coat protein (CP) deletion that causes defectiveness in cell-to-cell movement, together with a functional CP+3' non-translated region (ntr) transcript, in Nicotiana benthamiana leaf tissue. The structural parameters assessed were the length of sequence overlap, the distance between mutations and the degree of sequence similarity. The effects on the observed frequency of reconstitution and the composition of the recombination products were characterized. Application of four different type X intact PVX CP genes with variable composition allowed the estimation of the junction sites of precise homologous recombination. Although one template switch would have been sufficient for functional reconstitution, between one and seven template switches were observed. Use of PVX–GFP mutants with CP deletions of variable length resulted in a linear decrease of the reconstitution frequency. The critical length observed for homologous recombination was 20–50 nt. Reduction of the reconstitution frequency was obtained when a phylogenetically distant PVX type Bi CP gene was used. Finally, the prediction of CP and 3'-ntr RNA secondary structure demonstrated that recombination-junction sites were located mainly in regions of stem–loop structures, allowing the recombination observed to be categorized as similarity-assisted.

These authors contributed equally to this work.

Two supplementary figures showing CLUSTAL_X alignments are available with the online version of this paper.