Common barberry (Berberis vulgaris L.) is an alternate host for black stem rust of wheat, Puccinia graminis Pers. The shrub provides a means for the pathogen to reproduce after the winter and then spread to new cereal plants in the spring (Clark et al., 1986; Pierce, 1981). Wheat rust is a destructive fungal disease of cereals and grasses capable of causing major crop losses (Pierce, 1981; Roelfs, 1982). The rust multiplies on barberry and often develops new and more virulent races to which cultivars of cereal crops may have little or no resistance (Clark et al., 1986; Pierce, 1981; Roelfs, 1982). To control the emergence of black stem rust of wheat, the Canadian government has prohibited the importation of barberry species (MacLatchy, 1996). Japanese barberry (Berberis thunbergii DC.) is an ornamental shrub appreciated for its hardiness and attractiveness but falls under the Canadian government prohibition of importation and domestic movement of barberry species in Canada. Because Japanese barberry is considered generally immune to rust (Melander and Craigie, 1927), the Canadian government has developed a program permitting the importation, multiplication, and movement within Canada of only approved resistant cultivars of Japanese barberry (CFIA, 2002). Accurate identification of Japanese barberry cultivars, especially for the distinction of the 11 approved cultivars, is essential for the success of the program. In the application of the program, Japanese barberry cultivars are identified by government officials primarily based on morphology. However, variation in growth conditions or importation in a dormant state may influence or limit the number of morphologic characters in such a way that accurate identification becomes difficult.
DNA fingerprinting techniques have been proven to be reliable for cultivar identification because the markers are not influenced by environmental factors, life stage, or type of plant tissue analyzed. The choice of a fingerprinting method for cultivar identification depends mainly on the application and the time and cost related to method development. Semiarbitrary-primed polymerase chain reaction (PCR) methods were assessed for the selection of a DNA fingerprinting technique for the following reasons. 1) No sequencing data are required for their development; therefore, the time involved for the development of the method is shorter; this is especially useful for an ornamental crop that has not yet been extensively studied at the molecular level. 2) These fingerprinting methods are reasonably reproducible and generate a high number of polymorphisms per primer set, therefore allowing cultivar identification in a few PCR reactions. The amplified fragment length polymorphism (AFLP) technique was selected as the method of choice for Japanese barberry cultivar verification because it has proven to be reproducible and to generate highly polymorphic patterns (Vos et al., 1995).
AFLP is based on the selective amplification of restriction-digested DNA fragments of the genome of interest through the ligation of adaptors composed of the restriction enzyme site and selective nucleotides (Vos et al., 1995). Recently, AFLP has been used for the cultivar identification of various crops, including celery (Apium graveolens L.) (Li and Quiros, 2000), mango (Mangifera indica L.) (Kashkush et al., 2001), strawberry [Fragaria ×ananassa (Weston) Duchesne ex Rozier (pro sp.)] (Tyrka et al., 2002), apricot (Prunus armeniaca L.) (Geuna et al., 2003), and poinsettia (Euphorbia pulcherrima Willd. ex Klotzsch) (Parks and Moyer, 2004). In all of these studies, AFLP successfully and effectively differentiated cultivars, therefore allowing their identification. Furthermore, AFLP was used for the characterization of natural populations of Berberis species in Argentina, therefore demonstrating the applicability of the method for the Berberis genus (Bottini et al., 2002).
The main objective of this study was to use AFLP as a fingerprinting method for the identification of approved Japanese barberry cultivars. This was done by establishing a set of polymorphic AFLP bands used to score Japanese barberry samples submitted for cultivar verification. The established method was then verified using leaves or branches from Japanese barberry plants sampled in nurseries across Canada. Suspected mislabeled plant samples and samples from plants demonstrating unusual morphologic characters were tested as well. The AFLP method successfully confirmed the variety indicated on the label for both mature and dormant plants. Furthermore, the method allowed the identification of plants that were confirmed to be mislabeled by the exporter.
Arús, P., Aranzana, M.J. & Carbó, J. 2003 SSR and AFLP markers for germplasm evaluation and cultivar identification in peach Acta Hort. 606 34 40
Bottini, M.C.J., De Bustos, A., Jouve, N. & Poggio, L. 2002 AFLP characterization of natural populations of Berberis (Berberidaceae) in Patagonia, Argentina Plant Syst. Evol. 231 133 142
CFIA 2002 Plant protection import and domestic movement requirements for barberry (Berberis, Mahoberberis, Mahonia). Plant Health and Production Division Canadian Food Inspection Agency D 01-04 1 24
Clark, R.V., Seaman, W.L. & Martens, J.W. 1986 Puccinia graminis on barberry and oats in eastern Ontario from 1968 to 1983 Can. J. Plant Pathol. 8 193 200
Kashkush, K., Jinggui, F., Tomer, E., Hillel, J. & Lavi, U. 2001 Cultivar identification and genetic map of mango (Mangifera indica) Euphytica 122 129 136
Lanham, P.G. & Brennan, R.M. 1999 Genetic characterization of gooseberry (Ribes grossularia subgenus Grossularia) germplasm using RAPD, ISSR and AFLP markers J. Hort. Sci. Biotechnol. 74 361 366
Li, G. & Quiros, C.F. 2000 Use of amplified fragment length polymorphism markers for celery cultivar identification HortScience 35 726 728
MacLatchy, I.A. 1996 Berberis thunbergii (Japanese barberry) as a potential obstacle to the control of Puccinia graminis (Stem rust of wheat) Plant Health Risk Assessment Unit, Agriculture and Agri-Food Canada 3510-3-1B2
Montemurro, C., Simeone, R., Pasqualone, A., Ferrara, E. & Blanco, A. 2005 Genetic relationships and cultivar identification among 112 olive accessions using AFLP and SSR markers J. Hort. Sci. Biotechnol. 80 105 110
Parks, E.J. & Moyer, J.W. 2004 Evaluation of AFLP in poinsettia: Polymorphism selection, analysis, and cultivar identification J. Amer. Soc. Hort. Sci. 129 863 869
Tyrka, M., Dziadczyk, P. & Hortyński, J.A. 2002 Simplified AFLP procedure as a tool for identification of strawberry cultivars and advanced breeding lines Euphytica 125 273 280
Vos, P., Hogers, R., Bleeker, M., Reijans, M., van de Lee, T., Hornes, M., Frijters, A., Pot, J., Peleman, J., Kuiper, M. & Zabeau, M. 1995 AFLP; a new technique for DNA fingerprinting Nucleic Acids Res. 23 4407 4414