Single nucleotide polymorphisms (SNPs) are frequently observed in the genes as well as intergenic regions among related strains. And SNPs are effectively used for genotyping of various strains. In wheat, however, the nature of polyploidy prevents, to some extents, SNPs analysis by the ordinary methods. Critical point to carry out SNPs analysis in wheat is to assign the individual SNPs into each of three genomes, namely A, B and D. We developed an EST assembling method that can distinguish the contigs transcribed from each loci located on different parts of chromosomes. By way the method 116,232 EST sequences of Chinese Spring (CS) were assembled into 25,971 contigs. Out of 25,971 contigs containing more than 5 members were selected and classified into related gene groups by combination of BLAST and phrap methods. Single copy gene groups per genome were supplied for further analysis. The grouped contigs were aligned, and the SNPs positions were determined. The frequency of SNPs examined was estimated to be once per 144.9 bp. Pyrosequencing^TM method in combination with the nulli-tetrasomics series of CS was adopted to assign individual SNPs into each chromosome of hexaploid wheat. Most SNPs can be assigned into each chromosome, showing applicability of the Pyrosequencing method for SNPs analysis of hexaploid wheat.

S-2-4 K. Murai (Dept Biosci, Fukui Pref Univ; murai@fpu.ac.jp)
Functional genomics using EST data in wheat -- A case of MAPS box gene family --

To explore the gene expression underlying wheat life cycle, a large-scale analysis has been done on the cDNAs from various tissues at different growth stages in wheat. A set of about 70,000 expressed sequence tags (ESTs) was analyzed and grouped into about 26,000 independent clusters. Among these, 36 MADS box gene contigs were found. MADS box genes encode trascriptional regulators involved in diverse aspects of plant development. Based on the deduced amino acid sequences, the wheat MADS box genes were classified into seven groups. The largest group consists of 16 MADS box genes which show homology to AGL2 / SEP1 -like genes in Arabidopsis. It is well known that the sequence homology of MADS box genes is highly related to the functional homology. Four wheat MADS box gene clones, #3558, #4273, #5651 and #13055 of AGL2/SEP1-like group, were closely related to OsMADS1 in rice, and ZMM8 and ZMM14 in maize, which play an important role in floral meristem determination in spikelets. This suggests that these wheat MADS box genes may also be involved in conferring determinacy of floret identity.

S-2-5 K Kato (Obibiro Univ Agric & Vet Med)
Functional genomics combined with QTL analysis in wheat

To clarify the genie control of flowering time and seed dormancy in hexaploid wheat, we have applied comparative genetic mapping of QTLs and candidate genes across plant species including rice, maize and barley. We identified the genetic map positions of two candidate genes, tck2a on chromosome 5A and taVp1 on 3A. Comparative mapping showed that tck2a was syntenous to the rice CK2a or the heading date QTL Hd6. On the other hand, ta Vp1 was expected to be an orthologue of Vp1, the maize viviparous gene. However, our data did not support the direct relationship between candidate genes and QTLs. We will clarify the genie interaction between candidate genes and QTLs in future study. Alternatively, near isogenic lines carrying each QTL and/or chromosomal segment have been developed by marker-assisted selection using DNA markers. These plant materials will be profitable to characterize the functions of QTLs.

S-2-6 C. Nakamura (Grad Sch Sci & Technol, Kobe Univ)
Functional genomics approaches towards nucleus-cytoplasm interactions in wheat

The genetic system of the cytoplasmic genome (plasmon consisting of chloroplast and mitochondrial genomes) differs significantly from that of the nuclear genome in various aspects. The plasmon is semi-autonomous and yet can sustain its structural and functional integrity only through an intimate interaction with the resident nuclear genome. One important question directly related to the plasmon biology including nucleus-cytoplasm interaction and retrograde regulation of the nuclear gene expression is; if and how we can develop experimental systems that should enable us to carry out functional genomics research in wheat and its related species. The symposium talk is divided into three parts. First, several new trends in plant mitochondrial research are introduced, based on interesting reports in the 6th Int. Congress of Plant Mitochondria, July 9-14, 2002, Perth, Australia. Second, our recent research results and future perspectives of the cold responsive genes/proteins in wheat are reported. The genetic systems of low temperature and dehydration stress responses involving nuclear genes and proteins targeted to chloroplasts and mitochondria hopefully provide model systems for functional genomics study of the NC interaction. Third, a peculiar phenomenon of mitochondrial (mt) DNA heteroplasmy depicted as mixed presence of the maternal and paternal mtDNA copies in individual plants is addressed. Biological and evolutional significance of the observed mtDNA heteroplasmy remains largely unknown, but this system provides a new and unique tool for evolutional and functional genomics study in wheat and its related species.