| Rice (Oryza sativa L.) is not only a pivotal key cereal crop that provides the staple food for more than half of the world’s population as a source of daily carbon intake, but also a plant that has attracted broad interest in basic and applied research. The published of draft genome sequences of two rice cultivars to proclaim that the study of both the indica and japonica subspecies has entered into post-genomic era. The completed sequence variety Oryza sativa cv. Nipponbare as experimental material can be found in this paper. Seeds of this variety were treated with ethyl methane sulphonate (EMS). A total of2210materials were first screened by NIR from which208mutants were obtained. A second screening yielded73mutants that were all identified by the chemical experiment. The following results were observed in present experiment.1. A number of27mutants with appearance and cooking quality traits have been obtained:5chalky mutants were screened and the mutation frequency of chalkiness was2.26%o, followed by5amylose content (AC) mutants (2.26%o), both4gelatinization temperature (GT) and gel consistency (GC) mutants (1.81‰),2alkali spreading value (ASV) mutants (0.9%o), and7rapid visco analyzer (RVA) profile mutants (3.17%o).2. A number of61mutants of nutrition quality have been obtained:11protein content (PC) mutants were detected at a mutation frequency of4.98%o. Relative contents of17kinds of amino acid mutations, including7kinds of essential amino acid and10kinds of nonessential were identified. When the screening standard was set at10%, mutants of lysine and leucine (0.45‰), and valine (4.98‰) were screened, but no mutant was obtained for isoleucine, phenylalanine or threonine in essential amino acids. For nonessential amino acids, mutants of glutamic (0.45‰), arginine (3.17‰), alanine (2.71‰), serine (0.45‰), glycine (0.45‰), tyrosine (1.36‰), proline (2.26‰), and histidine (0.45‰) were obtained, but no mutant was obtained for aspartic, phenylalanine or threonine. With the screening standard for methionine and cysteine set at100%, the mutation frequencies of these two amino acids were2.26‰and4.98‰, respectively. Mutants of this preliminary library for rice quality mutants would play important role in the study of gene function and breeding for rice quality.3. A floury endosperm (flo6) mutant with chalky phenotype was used to constructed gene mapping population. The scanning electron microscopic results showed that the starch granules of flo6mutant were comprised round and loosely packed, some starch granules compounded by the film coating. Cooking and nutrition quality analysis indicated that the value of GC, GT or RVA profile of flo6mutant was lower than that of WT; and the thermodynamic parameters Tp andâ–³T1/2between WT and flo6mutant were different. PC of flo6mutant was higher than that of WT. According to the relative contents of17amino acids analyzed,9of them were different between WT and flo6mutant. Genetic analysis revealed that flo6mutation was controlled by a single recessive nuclear gene.4. The flo6locus was located on a500kb region by primary mapping. Map-based cloning and sequencing showed that there was a single nucleotide mutation (Gâ†'A) in LOC_Os01g44220.4.exon10, which resulted in the sequence of transcript protein of locus LOC_Os01g44220had been changed (Eâ†'K). This gene was annotated encodes glucose-1-phosphate adenylyltransferase large subunit in database, and it was named OsAPL2. The protein of this gene catalyzed the first step of starch synthesis in rice endosperm, and this reaction was considered to be the main limiting factors of rice starch synthesis. The tertiary structure had been changed because of the site mutation.5. The result of comparison of protein sequences showed that the amino acid residue mutant site was closed to the residues interacting with ADP-Glu of AGPase, and the original residue was Glu with negative charge mutated into Lys with positive charge, so the charge environment about the interact residues was changed by this mutation. Thus, the site mutation of protein sequence did not lead to the complete lost of gene function for OsAPL2, it just decreased its capability to catalyze or polymerize with others subunits. Based on the model of AGPase works, the content of AGPase complex which was composed by OsAPL2with other subunits was lower in the early stage of endosperm development, and the capability of the complex was also decreased because of the site mutant. Meanwhile, the enzymatic activity of AGPase complex which was composed by OsAPS1and OsAPL1was lower (only5%), the catalyzed activity was mostly supplied by the cytoplasm AGPase. As a result, at the early stage of endosperm development, the lack of ADP-G whcih was the substrate of starch synthesis, reduced the speed of starch synthesis, which was why the starch granules were loosen in the central section of grains of flo6mutant. At the later stage, the content of OsAPL2increased rapidly in a short time, so it was a compensation for the decrease of catalyzed capabilities that was induced by the site mutation. Under these conditions, the synthesis of ADP-G and starch returned to normal. Finally, the mutation phenotype at the interior of the endosperm was floury-white while the outer portion was normal has formed. |
PDF Full Text Request