Sequence and RFLP analysis of the elongation factor Tu gene used in differentiation and classification of phytoplasmas

• Published in: Microbiology (Society for General Microbiology) 1997-10, Vol.143 (10), p.3381-3389
• Main Authors: Schneider, Bernd, Gibb, Karen S
• Format: Article
• Language: English
• Subjects: Bacterial plant pathogens; Bacteriology; Base Sequence; Biological and medical sciences; Blotting, Southern; DNA Primers - genetics; DNA, Bacterial - genetics; Fundamental and applied biological sciences. Psychology; Gene Amplification; Genes, Bacterial; Genetics; Microbiology; Molecular Sequence Data; Peptide Elongation Factor Tu - genetics; Phylogeny; Phytopathology. Animal pests. Plant and forest protection; phytoplasma; Plants - microbiology; Polymerase Chain Reaction; Polymorphism, Restriction Fragment Length; Systematics. Structure, properties and multiplication. Genetics; Tenericutes - classification; Tenericutes - genetics
• ISSN: 1350-0872; 1465-2080
• DOI: 10.1099/00221287-143-10-3381

Faculty of Science, Northern Territory University, Darwin 0909, Australia Biologische Bundesanstalt für Land- und Forstwirtschaft Erich Seemüller, Institut für Pflanzenschutz im Obstbau, 69221 Dossenheim, Germany 1 Author for correspondence: Bernd Schneider. Tel: [phone]. Fax: [phone]. e-mail: [e-mail] ABSTRACT Primers designed from sequences of the gene encoding the elongation factor Tu ( tuf gene) of several culturabie mollicutes amplified most of the tuf gene from phytoplasmas of the aster yellows, stolbur and X-disease groups. About 85% of the tuf gene from two aster yellows strains and a tomato stolbur phytoplasma was sequenced. The nucleotide sequence similarity between these related phytoplasmas was between 87.8 and 97.0%, whereas the homology with other mollicutes was 66.3-72.7%. The similarity of the deduced amino acid sequence was significantly higher, ranging from 96.0 to 99.4% among the phytoplasmas and 78.5% to 83.3% between phytoplasmas and the culturabie mollicutes examined. From the nucleotide sequences of the phytoplasma strains, two pairs of primers were designed; one amplified the phytoplasmas of most phylogenetic groups that were established, the other was specific for the aster yellows and stolbur groups. The phytoplasmas of the various groups that were amplified could be distinguished by RFLP analysis using Sau 3AI, Alu I and Hpa ll. The aster yellows group could be divided into five Sau 3AI RFLP groups. These results showed that the tuf gene has the potential to be used to differentiate and classify phytoplasmas. Southern blot analysis revealed that the tuf gene is present as a single copy. Keywords: phytoplasmas, differentiation, classification, elongation factor, PCR