ISHS Acta Horticulturae 472: XVII International Symposium Virus and Virus-Like Diseases of Temperate Fruit Crops
IC-PCR AMPLIFICATION OF APPLE STEM GROOVING VIRUS ISOLATES AND COMPARISON OF POLYMERASE AND COAT PROTEIN GENE SEQUENCES |
| Authors: | S.J. Crossley, V. Jacobi, A.N. Adams |
| Keywords: | capillovirus, citrus tatter leaf virus |
| Abstract:
Apple stem grooving virus (ASGV) is a capillovirus which infects a wide range of apple and pear cultivars. Published primers have been used in immuno-capture PCR (IC-PCR) experiments to amplify part of the coat protein region of two UK isolates of ASGV. The sequence data obtained has been compared with published sequence data from a Japanese isolate of ASGV and two isolates of citrus tatter leaf virus (CTLV). Using these comparisons five new primers were designed to conserved areas. One new primer pair have been used to amplify a 345bp coat protein product from samples of 10 apple and pear varieties from the National Fruit Tree Collection, Brogdale, UK. Following sap inoculation of these viruses into Chenopodium quinoa, a second primer pair were used to amplify a much larger coat protein product (765bp) from the ten isolates for sequencing purposes. In addition, the isolates were amplified with primers which gave a 756bp product from the polymerase gene region and this was also sequenced. Sequence data were generated from PCR products directly and from cloned PCR products. Sequence information from PCR products revealed that some samples contain mixed populations of virus and that some of these differences produce changes in the deduced amino acid sequence. Multiple sequence alignments of the regions amplified show that the coat protein gene is more highly conserved than the polymerase gene and therefore a better choice in the design of diagnostic primers. Furthermore, the sequence data for the coat protein region falls into two distinct groups, one containing the Japanese pear isolates and the other containing the European apple isolates. | |
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