Conversion of AFLP markers linked to the S^h allele at the S locus in buckwheat to a simple PCR based marker form

Mio Nagano¹, Jotaro Aii¹, Masako Kuroda¹, Clayton Campbell², and Taiji Adachi³.

¹ Applied Genetics and Biotechnology Division, Faculty of Agriculture, Miyazaki University, Gakuen Kibanadai Nishi 1-1, Miyazaki 889-2192, Japan ² Kade Research Ltd., Morden, Manitoba, Canada R6M 1E9 ³ Laboratory of Plant Genes and Physiology, Graduate School of Agriculture and Biological Sciences, Osaka Prefecture University, 1-1 Gakuen-cho, Sakai, Osaka, 599-8531, Japan

Mio Nagano, Jotaro Aii, Masako Kuroda, Clayton Campbell, and Taiji Adachi. 2001. Conversion of AFLP markers linked to the S^h allele at the S locus in buckwheat to a simple PCR based marker form. Plant Biotech. 18, 191-196.

Abstract: Using the technique of amplified restriction fragment length polymorphism (AFLP) analysis approximately 500 polymorphic loci were screened on the bulked segregant pools from F₂ progeny of the cross between Fagopyrum esculentum (pin) and F. homotropicum. The objective was to find those markers with tight linkage to the buckwheat homostylar locus, concerned with self-incompatibility. Analysis of 123 F₂ plants identified nine markers that show no recombination in 36 recessive homozygous plants. In the nine markers, two (N2 and N7) were confirmed to derive from a single region. The N2 sequence was present only in F. homotropicum and was absent in common buckwheat, F. esculentum. Nucleotide sequence information from each flanking region of the two single locus markers was used to design region-specific primers for PCR amplification. The N2 region-specific primer amplified a single fragment in F. homotropicum #1 but not in common buckwheat, F. esculentum #284 (pin). Whereas the N7 AFLP marker was converted into a co-dominant marker for both parents. However, the N7 marker showed size polymorphism between the parent lines. These markers can be utilized for fine mapping of the S^h allele in buckwheat and for positional cloning of the gene.

Key words: Fagopyrum esculentum, F. homotropicum, buckwheat, AFLP markers, PCR marker