Ribosomal DNA intergenic spacer sequence in foxtail millet, Setaria italica (L.) P. Beauv. and its characterization and application to typing of foxtail millet landraces.

摘要

Ribosomal DNA (rDNA) in plants, as in animals, is arranged in tandem arrays and contains genes for 25 S, 18 S and 5.8 S rRNAs. These genes are highly conserved among species, but spacer regions such as internal transcribed spacers (ITS) and intergenic spacers (IGS) of the gene are less conserved than the genes themselves. Spacer regions are often used for constructing phylogenetic trees among species (Buckler and Holtsford 1996; Yasui and Ohnishi 1998; Sallares and Brown 2004) and detecting intraspecific polymorphisms (Saghai-Maroof et al. 1984; Cordesse et al. 1990; Sano and Sano 1990; Fukunaga et al. 1997; Polanco and Pérezde la Vega 1997). In particular, IGS has highly variable length, even within species or among cultivars, because it contains subrepeats which vary in repeat number (Rogers and Bendich 1987). The upstream of the IGS sequence is so divergent between distantly related species even in the same family (Cordesse et al. 1993; Polanco and Pérezde la Vega 1994; Da Rocha and Bertrand 1995; Fernández et al. 2000). However, it was reported that the sequences of external transcribed spacer (ETS) at the downstream part of the IGS are less divergent and this region became a tool for studying phylogeny among closely related species (Baldwin and Markos 1998; Linder et al. 2000; Sallares and Brown 2004).Foxtail millet (Setaria italica (L.) P. Beauv.), belonging to the subfamily Panicoideae, is an ancient domesticated cereal from the Old World. Molecular studies have been carried out to address the diversity of this crop (rDNA: Fukunaga et al. 1997; Schontz and Rether 1998; 5S rDNA: Benabdelmouna et al. 2001; RAPD: Li et al. 1998; Schontz and Rether 1999; nuclear RFLP: Fukunaga et al. 2002b; AFLP: Le Thierry d'Ennequin et al. 2000; mitochondrial DNA: Fukunaga and Kato 2003; waxy gene: Fukunaga et al. 2002a) because it has played an important role in the history of Old World agriculture. It remains important in China (Li and Wu 1996). Recently, an RFLP linkage map has been constructed for foxtail millet (Devos et al. 1998; Wang et al. 1998). This crop has also become an interesting material in development and evolution of grass species (Doust and Kellogg 2002; Doust et al. 2004, 2005). In Japan, there are two collection sets of foxtail millet germ-plasms, one at Kyoto University consisting of about 1000 accessions and the other at Kagoshima University of about 600 accessions.For foxtail millet, Fukunaga et al. (1997) and Schontz and Rether (1998) investigated rDNA polymorphisms. Those studies revealed genetic variation and geographical distribution patterns of each variant. Both of them used genomic Southern hybridization to detect three major types, which differ in length (ca 300 bp) and a BamHI site in repeat units. Types I, II and III by Fukunaga et al. (1997) correspond to types A, B2 and B1 by Schontz and Rether (1998), respectively. However, these two reports reached different conclusions regarding geographical distribution patterns. Fukunaga et al. (1997) concluded that type I is mainly distributed in the temperate zone while types II and III are mainly distributed in the Taiwan-Philippines (Nansei Islands of Japan, Taiwan and Batan Islands of the Philippines) and India, respectively. On the other hand, Schontz and Rether (1998) concluded that type I is mainly distributed in Europe while types II and III are mainly distributed in southern and northern Asia, respectively. Genomic Southern hybridization is an onerous procedure for accessing a large number of samples. For the maintenance and characterization of the germ-plasms, development of PCR-based molecular markers is essential.In the present study we determined and characterized the sequence of the rDNA IGS. We also developed PCR-based typing of rDNA according to the sequence information and added some information on rDNA polymorphism and geographical distribution by examining the collection set of Kagoshima University.