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M. Murata (Res Inst Bioresour, Okayama Univ; mmura@rib.okayama-u.ac.jp)
Construction of chromosome-specific DNA libraries by laser-microdissection

We are constructing chromosome arm-specific DNA libraries in wheat by using laser-microdissettion. Telocentric chromosomes in ditelosomic lines were chosen as targets, scratched off and picked up with glass needles adjusted to a micromanipulator. The microdissected chromosomes were then harvested into a PCR tube, and their DNA was amplified using DOP-PCR. In order to evaluate the efficiency of our nicrodissection procedure, we attempted to amplify DNA from only one telocentric chromosome, and compared with the results from two, and four microdissected fragments. In all cases, distinct DNA amplification could he observed. Sequencing analysis revealed that a relatively high proportion of low-copy sequences are involved in the PCR fragments. This suggests the present microdissection procedure is effective in creating painting probes and in generating region-specific DNA markers.

T. Wako1, A. Houben2 , R. Furushima-Shimogawara3, B. M. Turner 4 and K. Fukui5
(1Natl Inst Agrobiol Resour, 2Adelaide Univ, 3Tokyo Sci Univ, 4Dept Medicine, Univ Birmingham, 5Dept Biotech, Graduate School Engineer, Osaka Univ)
Three dimensional analysis of histone acetylation and phosphorylation on mitotic chromosomes in cereals

Histone acetylation and phosphorylation affect chromatin conformation and regulate various cellular function. Changes in acetylation of H4 at lysine 5 (K5) and 16 (K16), and phosphorylation of H3 at serine 10 (S10) during mitosis have been examined by three-dimensional microscopy. Telomeric region was enriched acetylated H4 at K16 throughout mitosis. Centromeric region was enriched acetylated H4 at K5 when chromosomes were decondensed and phosphorylated H3 at S10 when chromosomes were condensed. Nucleolar organizing regions were acetylated at K5 between prophase to anaphase. We propose that H4 acetylation and H3 phosphorylation define functional chromatin domains throughout the cell cycle.


2. Symsium on recent topics and prospects in wheat and barley breeding (Organizer: T. R. Endo)

T. Nakamura1*, P. Vrinten1 and K. Hayakawa2 (1Dept Crop Breed, Tohoku Natl Agr Expt Stan, 2 Cereal Res Cent, Nisshin Flour Milling Co; *tnaka@tnaes.affrc.go.jp)
Waxy Wheat: Production, properties and mutations

Partial waxy lines were used to produce both hexaploid (common) and tetraploid (durum) waxy wheat lines. The starch of these lines lacks amylose, and this change in composition drastically alters starch properties. Our primary application tests indicate that blending waxy and regular wheat flour improves texture and resistance to starch retrogradation in products such as noodles, bread and Chinese dumplings. The mutations in a hexaploid waxy wheat line produced by our group were analyzed and all three alleles were found to carry deletions. Although the three Wx genes in waxy wheat are nonfunctional, amylose was present in the pericarp starch of waxy wheat, and a second GBSS isozyme (GBSSII) was detected in pericarp starch. A GBSSII cDNA was isolated, and expression analysis indicated GBSSII mRNA was present in leaf, culm, and pericarp tissue, but was not detected in endosperm tissue.

Ali Masoudi-Nejad (Lab Plant Genetics, Div Applied Biosci, Kyoto Univ; amasoudin@kais.kyoto-u.ac.jp)
Wheat storage protein: present and future

Wheat seed storage proteins have been studied extensively for their pivotal role in determining nutritional and bread-making quality of flour. Because of their high proline and glutamine content they are called prolamin. Wheat prolamins are synthesized on the endoplasmic reticulum in the developing endosperm. They are classified as glutenin and gliadins, which are controlled by the genes located on the long arms of the homoeologous group 1 chromosomes (Glu-1) and those on the short arms of groups 1 and 6 chromosomes (Gli-1 and Gli-2), respectively. The last few decades witnessed a rapid advance in our knowledge on the wheat storage proteins, especially through the progress in the basic sciences like biochemistry and molecular biology. Numerous gene sequences coding for glutenin and gliadin have been isolated, cloned and characterized. This has allowed wore deep understanding of their structure, function and evolution. In this talk I review the past and our current knowledge about glutenin arid gliadin from different point of view incorporating the results of recent developments in molecular genetics and biochemistry. I also discuss works undergoing in our laboratory on sequencing of an omega-gliadin gene and its deletion mapping.

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