Tomoaki Taira

The isolation and characterization of a waxy mutant of diploid wheat (Triticum monococcum L.)

A waxy mutant of diploid wheat (Triticum monococcum L.) was isolated by screening M(3) seeds derived from 1% ethyl methanesulfonate (EMS) mutagenized materials with KI-I(2) staining of endosperm starch. This mutant was controlled by a single waxy gene that completely lacked amylose, the 59-kDa waxy protein and the granule-bound starch synthase I (GBSS I) activity in the endosperm. Moreover, the T. monococcum exhibited the gene dosage effects in terms of amylose content, the amount of waxy protein and the GBSS activity. The chain length distribution of amylopectin of the waxy mutant endosperm starch slightly differed from that of the wild type; the long chains of DP>==19 were more enriched in the waxy amylopectin than those in the wild type. The mutant can be an important material for studies on starch metabolism and breeding in Triticum species.

A 56-kDa protein is a novel granule-bound starch synthase existing in the pericarps, aleurone layers, and embryos of immature seed in diploid wheat (Triticum monococcum L.)

Abstract. A novel 56-kDa granule-bound starch synthase (GBSS; NDPglucose-starch glucosyltransferase, EC 2.4.1.21) responsible for amylose synthesis was found in the pericarps, aleurone layers and embryos of immature diploid wheat (Triticum monococcum L.). The GBSS and other proteins bound to starch granules of various tissues of immature normal and waxy diploid wheat seeds were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and their activities were examined. In the waxy mutant, the waxy protein (59.5 kDa, GBSSI) was absent, but amylose and GBSS activity were evident in all tissues except the endosperm. Of the proteins bound to starch granules, only the 56-kDa protein was associated with the presence of amylose and GBSS activities in the pericarps, aleurone layers and embryos. Mutations at the waxy locus did not affect the 56-kDa protein in these tissues. Changes in the amount of 56-kDa protein during the course of seed development, and the distribution of the 56-kDa protein in each tissue of immature seeds were quite different from those of the waxy protein. On the other hand, the N-terminal amino acid sequence of the 56-kDa protein had a 40–50% similarity to GBSSI of some other plant species and was antigenically related to the waxy protein. These results strongly suggest that the 56-kDa protein in diploid wheat is a GBSSI class enzyme and, hence, an isoform of the waxy protein. The waxy protein and 56-kDa protein, however, are expressed in different seed tissues and at different stages of seed development.

Change of Ascorbic Acid Level after Grafting of Tomato Seedlings

Grafting is an easy way to produce a new seedling, which can tolerate against various stresses. During the acclimation after grafting, however, the seedlings still suffer a severe water stress. It is well known that water stress produces active oxygen to oxidize ascorbic acid. The concentration of ascorbic acid in the leaves was analyzed by HPLC equipped with an electrochemical detector. The column used was SP-120-5-ODS-BP (DAISO, JAPAN) and elution was performed with 0.1 ᴍ phosphate buffer, pH 3.0. After grafting the seedlings were acclimated under a 6-hr light/dark regimen. The content of ascorbic acid increased gradually during 2 days compared with control. The ascorbate peroxidase showed about constant activity, so the increase of ascorbic acid may be due to its requirement to cure the grafting