ISHS Acta Horticulturae 521: XXV International Horticultural Congress, Part 11: Application of Biotechnology and Molecular Biology and Breeding - Gene Expression and Molecular Breeding, Genome Analysis.
INVESTIGATION OF MICROSATELLITE PROFILING FOR DUS TESTING IN CHRYSANTHEMUM |
| Authors: | J.A. Jackson, D. Matthews, W. Cooper, R.J. Cooke |
| Keywords: | 5'-anchored inter-SSR PCR, Chrysanthemum, DUS, ISTR, microsatellites |
| Abstract:
DNA fingerprinting techniques based on PCR amplification of microsatellite and Copia like sequences were investigated for their utility in the Chrysanthemum DUS process. Microsatellite primers with 3 and 7 bp 5' degenerate anchors and either 6, 12 or 18 tandem repeats were compared with the retrotransposon primers of Rhode (1996). Dendrograms were constructed from the profile data and compared with the known lineages of the cultivars investigated. A primer comprising a 7 bp 5' degenerate anchor (KKVRVRV) and a CT18 repeat most accurately recreated the known relationships between the cultivars. PCR products generated by CT-6, 12 and 18 primers were cloned and sequenced and all inserts were found to contain at least two microsatellites. The average length of the microsatellites and the variation in the number of tandem repeats increased as the length of the primer increased. Sequence data from the inter-microsatellite region was used to design specific primers. These were used in conjunction with the anchored primers for amplifying single loci. The benefits of these approaches is discussed. | |
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