Characterization of putative membrane protein genes of the ‘Candidatus Phytoplasma asteris’, chrysanthemum yellows isolate

Author:

Galetto Luciana123,Fletcher Jacqueline123,Bosco Domenico123,Turina Massimo123,Wayadande Astri123,Marzachì Cristina123

Affiliation:

1. Università degli Studi di Torino, Facoltà di Agraria, Di.Va.P.R.A, Entomologia e Zoologia Applicate all’Ambiente “Carlo Vidano”, Via L. Da Vinci 44, Grugliasco (TO) 10095, Italy.

2. Istituto di Virologia Vegetale, CNR, Strada delle Cacce 73, Torino 10135, Italy.

3. Oklahoma State University, Department of Entomology and Plant Pathology, 127 Noble Research Center, Stillwater, OK 74078, USA.

Abstract

To characterize potentially important surface-exposed proteins of the phytoplasma causing chrysanthemum yellows (CY), new primers were designed based on the conserved regions of 3 membrane protein genes of the completely sequenced onion yellows and aster yellows witches’ broom phytoplasmas and were used to amplify CY DNA. The CY genes secY, amp, and artI, encoding the protein translocase subunit SecY, the antigenic membrane protein Amp and the arginine transporter ArtI, respectively, were cloned and completely sequenced. Alignment of CY-specific secY sequences with the corresponding genes of other phytoplasmas confirmed the 16S rDNA-based classification, while amp sequences were highly variable within the ‘Candidatus Phytoplasma asteris’. Five CY partial sequences were cloned into the pRSetC expression vector, and 3 of the encoded protein fragments (Amp 64/651, Amp 64/224, ArtI 131/512) were expressed as fusion antigens for the production of CY-specific polyclonal antibodies (A416 against Amp 64/224; A407 against ArtI 131/512). A416 recognized, in Western blots, the full-length Amp from CY-infected plants (periwinkle, daisy) and insect vectors ( Euscelidius variegatus , Macrosteles quadripunctulatus ). A416 also reacted to European aster yellows, to primula yellows phytoplasmas, to northern Italian strains of ‘Ca. Phytoplasma asteris’ from lettuce and gladiolus, but it did not react to American aster yellows phytoplasma.

Publisher

Canadian Science Publishing

Subject

Genetics,Molecular Biology,Applied Microbiology and Biotechnology,General Medicine,Immunology,Microbiology

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