Interspecific rice hybrid of Oryza sativa x Oryza nivara reveals a significant increase in seed protein content

Wild species offer a potential reservoir of genetic variation for crop improvement. Besides the valuable genes for disease resistance that the wild species have provided for rice improvement, recent studies have shown that these wild species could also provide favorable alleles for the improvement of yield and yield-related traits. The present study reports yet another potential of wild relatives of rice, which involves the improvement of seed protein content. A significant increase in seed protein content was observed in an interspecific hybrid between Oryza sativa ssp. indica and the wild species Oryza nivara. The hybrid showed a protein content of 12.4%, which was 28 and 18.2% higher than those of the parents O. nivara and IR 64, respectively. The increase in protein content was dependent on the genetic background of the rice variety used in the hybridization. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of seed storage proteins demonstrated that a significant increase in prolamins and glutelins was mainly responsible for the elevated protein content of the hybrid. Amino acid analysis of seed proteins revealed that the hybrid had net gains of 19.5% in lysine and 19.4% in threonine over the O. nivara parent on a seed dry weight basis. Molecular analysis indicated that the increase in protein content of the hybrid was not a result of chromosomal rearrangements or transposable element activation, at least in the chromosomal regions containing seed storage protein genes. A preliminary genetic analysis of the F 2 segregating population showed that the inheritance of the increased protein content was polygenic in nature. The development of this interspecific hybrid offers a great potential for selecting new rice cultivars that combine the high yield and superior cooking quality of IR 64 with improved seed protein content.     

Hybridization of AA genome rice species from Asia and Australia II. Meiotic analysis of Oryza meridionalis and its hybrids

To determine the genomic constitution of Oryza meridionalis Ng (2n=2x=24) and to estimate genomic affinity between Asian and Australian wild species of rice containing the AA genomes, chromosome pairing was analyzed at metaphase I in O. meridionalis, O. rufipogon Griff. (2n=2x=24), O. nivara Sharma et Shastry (2n=2x=24), O. sativa f. spontanea Rosch, and their artifical hybrids. The Oryza parental species and their F1 hybrids showed normal meiosis, but slightly reduced chiasma fequency was observed in the hybrids. It is concluded from the cytological analysis that (i) the Australian O. meridionalis contains the AA genome which has very high affinity to that in the Asian AA genome wild species of rice; (ii) chromosome structual changes, such as inversions and reciprocal translocations, have occurred in the genomes of the different species studied.     

Amino acid variation in the 10 kDa Oryza prolamin seed storage protein

The deduced amino acid variability for the 10 kDa prolamin was determined for 16 Oryza species, both cultivated (rice) and wild. Prolamin, a seed storage protein and site of nitrogen and sulfur accumulation, is sequestered in the subaleurone layer of the starchy endosperm for use during seedling germination. The 10 kDa prolamin amino acid distribution for the cultivated species (O. sativa and O. glaberrima) was determined and compared to those of wild and, hitherto unknown, noncultivated Oryza species. Four wild species (O. granulata, O. australiensis, O. brachyantha, and O. meyeriana) exhibited the greatest residue heterogeneity in both the signal and mature peptide regions. A breakdown of the essential amino acid variance among three Central/South American and one African endemic wild species is also presented and compared with those of rice.     

Hybridization of AA genome rice species from Asia and Australia I. Crosses and development of hybrids

Interspecific and intraspecific crosses of rice species were made involving the AA genome wild species Oryza meridionalis Ng from Australia, O. nivara Sharma et Shastry, O. rufipogon Griff, the weedy type O. sativa f. spontanea Rosch., and the cultigen O. sativa L. from Asia. Seed set and the number of hybrids obtained from both interspecific and intraspecific crosses were low and no significant differences were observed between the interspecific and intraspecific crosses. In the combination O. meridionalis × O. nivara and O. meridionalis × O. rufipogon, considerable differences in reciprocal crosses were observed, whereas higher seed set and more hybrids were obtained when O. meridionalis was used as the female parent. Pollen stainability and seed fertility of hybrids were very low, averaging between 1% and 9% in both interspecific and intraspecific combinations, except in the O. nivara intraspecific cross of 105386 × 106111, whose hybrid showed 83.0% pollen stainabi lity and 66.5% seed fertility.     

Identification of introgression lines of Oryza glaberrima Steud. with high mineral content in grains

Mineral concentrations in cereals are crucial for human health, especially for people who consume cereals as a subsistence diet. Although rice (Oryza sativa L.) is one of most important staple crops in the world, starch constitutes a large portion of its grains, and mineral concentrations are lower than in other staple crops. One solution is the improvement of mineral concentrations in rice grains through biofortification, and this approach requires investigation of genetic resources that confer high mineral concentrations to rice grains. In the present study, we performed 2 years of field experiments, in 2012 and 2013, and determined sodium (Na), magnesium (Mg), phosphorus (P), potassium (K), calcium (Ca), boron (B), manganese (Mn), iron (Fe), copper (Cu), zinc (Zn) and molybdenum (Mo) concentrations in the rice grains of 40 introgression lines derived from a cross between a japonica cultivar, Taichung 65 (T65), and African rice, Oryza glaberrima Steud. Substantial variation in mineral concentrations was observed among the 40 introgression lines. We selected several elite lines that had significantly higher concentrations of minerals, including Fe, Cu, Zn, Mo, Mg, P and Ca than the cultivated rice T65. These lines could be novel potential materials for breeding programs for biofortification and provide us with positional information for the candidate loci in the O. glaberrima genome responsible for high mineral concentrations in rice grains.     

Identification and variation of glutelin alpha polypeptides in the genus Oryza assessed by two-dimensional electrophoresis and step-by-step immunodetection

To obtain fundamental information for nutritional improvement of rice (Oryza sativa) seed proteins, the alpha polypeptides of the major storage protein glutelin varied over the genus Oryza were qualitatively and quantitatively characterized with unique methods. The polypeptides were maximally separated by two-dimensional electrophoresis (2D-PAGE) composed of nonequilibrium pH gradient gel electrophoresis (NEPHGE) and higher temperature SDS-PAGE. Then the subunit for each polypeptide spot was identified with the sequential immunodetection called a step-by-step detection method, making use of highly subunit-specific antibodies. The comparative analysis showed considerable variation in the accumulation level of A-type and B-type glutelin subunits and found unknown glutelin subunits that were unable to be identified with the antibodies used. Wild species accumulating a high amount of lysine-rich B-type glutelin subunits and unknown unique subunits were identified as they might play a crucial role in nutritional quality improvement of the cultivated rice.     

Grain qualities and their genetic derivation of 7 new rice for Africa (NERICA) varieties

NERICA are interspecific rice varieties from crossing between the high-yielding Asian rice ( Oryza sativa spp. Japonica) with locally adapted African rice ( Oryza glaberrima). In this study, we analyzed grain qualities of 7 NERICA varieties (NERICA 1 to 7) and genetic derivation of quality-related genes. Quality analyses of NERICA grains showed that 7 NERICA varieties were clearly classified into two groups based on the difference of amylose content, and the difference influenced the pasting and physical properties of grains. Genetic analysis of the gene encoding granule-bound starch synthase I (GBSSI), which is known as a key enzyme on amylose synthesis in rice grain, revealed that varieties with higher amylose content ( approximately 29%) have the gene derived from O. glaberrima parent, and group 2 with lower amylose content ( approximately 22%) have the gene from O. sativa parent. These results indicated that the difference in amylose content as well as grain properties among 7 NERICA varieties is mainly determined by the genetic derivation of GBSSI. Further genetic analysis of starch synthesis-related genes suggested that the genetic derivation of SSIIa also influences the chain length of amylopectin in 7 NERICA varieties.     

Monophyletic vs. polyphyletic origin of the crops on which agriculture was founded in the Near East

The following comparisons between crops and their closely related wild relatives provide clues for discriminating between monophyletic and polyphyletic origins under domestication: (i) Presence or absence of patterns indicative of founder effects in the cultivated genepool, compared to the amount of variation present in its wild progenitor. (ii) Uniformity or lack of uniformity (within a crop) in genes governing principal domestication traits (traits that were automatically selected for once the wild progenitor was introduced into cultivation). (iii) Species diversity: The number of closely related (congeneric) wild species with similar potential for domestication, native to the area under consideration; and how many of them entered cultivation. The present paper evaluates the information available on the eight crops that founded Neolithic agriculture in the Near East; and arrives at the conclusion that emmer wheat Triticum turgidum L. subsp. dicoccum Schúbler, einkorn wheat T. monococcum L., pea Pisum sativum L., and lentil Lens culinaris Medik. were very likely taken into cultivation only once or – at most – a very few times. Also chickpea Cicer arietinum L., bitter vetch Vicia ervilia (L.) Willd., and flax Linum usitatissimum L. seem to have been domesticated in a similar way, but the evidence concerning them is much scarcer. Only for barley Hordeum vulgare L. are there indications that it has been domesticated more than once – but again only a very few times.     

水稻野生型愈伤组织中耐盐细胞频率的影响因素

利用组织培养技术研究愈伤组织的倍性、品种、外植体对野生型愈伤组织中耐盐细胞频率的影响。发现 :( 1)单倍体和二倍体愈伤组织的耐盐细胞频率比较接近 ,四倍体愈伤组织的耐盐细胞频率一般比单倍体和二倍体愈伤组织的小 ;( 2 )不同品种的耐盐细胞频率差异大 ;( 3)二倍体植株幼穗培养产生的二倍体愈伤组织和四倍体植株花药培养产生的二倍体愈伤组织的耐盐细胞频率没有显著的差异     

Activity and composition of the methanogenic archaeal community in soil vegetated with wild versus cultivated rice

Wild rice (Oryza rufipogon) is a problematic weed in fields of cultivated rice (Oryza sativa). We hypothesized that the composition and/or the activity of the methanogenic microbial communities might be different in soil grown with cultivated versus wild rice. We used samples from Hainan, China, where wild rice grew on a field adjacent to cultivated rice. The composition of the methanogenic archaeal community was analyzed in samples of rice soil by targeting the 16S rRNA gene. Analysis of the terminal restriction fragment length polymorphism (T-RFLP) showed similar patterns in soil from wild versus cultivated rice. Sequences of archaeal 16S rRNA genes also showed similar composition in soil from wild versus cultivated rice, revealing the presence of Methanosarcinaceae, Methanosaetaceae, Methanobacteriales, Methanocellales (Rice Cluster I), Rice Cluster II, Crenarchaeota Group I.3 and Crenarchaeota Group I.1b. Incubation of soil samples under anoxic conditions generally resulted in vigorous CH₄ production after a lag phase of 7-8 days. Production of CH₄ was partially inhibited by methyl fluoride, a specific inhibitor of acetoclastic methanogenesis, resulting in nearly stoichiometric accumulation of acetate. CO₂ was produced without lag phase. The δ¹³C of the produced CO₂ was slightly lower in soil grown with cultivated rice versus wild rice, reflecting the δ¹³C of organic matter, which was about −29‰ for cultivated rice soil and about −24‰ for wild rice soil. The δ¹³C of the produced CH₄ and the acetate that accumulated in the presence of CH₃F was much more negative in cultivated versus wild rice soil, mainly since the isotopic fractionation factors for hydrogenotrophic methanogenesis were higher for soil from cultivated rice (α = 1.054) versus wild rice (α = 1.039). However, the percentage contribution of hydrogenotrophic methanogenesis to total CH₄ production was similar in both soils (27-35%). In conclusion, although the two soils exhibited different δ¹³C values of soil organic matter and derived products, they were similar with respect to rates and composition of the methanogenic communities.     

水稻OsHsfA7基因RNA干扰载体的构建及遗传转化研究

为研究水稻热激转录因子基因OsHsfA7的功能及其在水稻耐热育种方面的应用价值,本文通过构建水稻OsHsfA7基因RNA干扰载体获得功能抑制变异材料,从反向遗传学进行基因的功能分析。扩增OsHsfA7c DNA3'编码区470bp片段,分别以反向和正向插入到中间载体pBSK连接的GUS片段两侧,然后将得到的RNA干扰片段克隆到改造的植物表达载体p1301M,构建以CaMV35S启动子驱动表达的水稻OsHsfA7基因RNA干扰表达载体。将该载体转入根癌农杆菌EHA105后,采用农杆菌介导法进行了水稻日本晴品种的遗传转化,获得了23株具有潮霉素抗性的转基因植株,其中12株经GUS染色呈蓝色并且DNA检测10株已插入目的片段,RNA检测其中6株OsHsfA7基因的表达水平下降甚至未检出。结果说明本研究构建的OsHsfA7基因RNA干扰载体对该基因表达沉默是有效的。     

普通野生稻稻米加工品质和外观品质性状QTL定位

本研究利用一套以籼稻品种“特青”为遗传背景的云南元江普通野生稻（Oryza rufipogon Griff.）渗入系为材料，采用单标记回归分析和渗入片段叠代法，对出糙率、整精米率、垩白粒率、垩白度、长宽比等5个品质性状的QTL进行了分析，初步定位了16个QTL，有10个QTL来自野生稻的等位基因能改良群体的品质性状。在第5染色体RM289附近检测到了同时影响长宽比、垩白粒率QTL，来自野生稻的等位基因能增加长宽比、降低垩白粒率，贡献率也较高。在第8染色体RM152附近检测到降低垩白粒率和垩白度的QTL，其贡献率分别为14％和9%。本研究结果不仅为品质性状分子标记辅助选择提供参考，而且充分显示了利用野生稻的优异基因改良栽培稻品质性状的巨大潜力。     

一个水稻脆性突变体的遗传分析与基因定位

化学诱变处理籼稻品种E532,M3代中筛选出茎、叶均较脆的突变体,定名fr(fragile rice)。遗传分析表明,该脆性突变体受两对隐性基因叠加控制。以fr突变体与粳稻02428杂交的F2代群体为基因定位群体,利用SSR分子标记将fr突变位点分别定位在1号染色体上的SSR标记RM302和RM212外侧,遗传距离分别为11.89cm和15.19cm;7号染色体在RM11和RM234之间,遗传距离分别为9.36cm和10.01cm。这些结果为进一步研究脆性突变体及其基因精细定位打下基础。     

云南元江野生稻抽穗开花期耐热QTL定位

利用以籼稻(Oryza sativa ssp. indica) 品种特青为遗传背景的云南元江普通野生稻（O. rufipogon Griff.）(简称“元江普野”)渗入系为材料,调查温室高温胁迫条件下野生稻渗入系和受体亲本特青的结实率,采用单标记回归分析法,共检测到4个抽穗开花期耐热性相关的QTL,分别位于第1、3、8和10染色体。其中位于第1和3染色体上的2个QTL（qHT1和qHT3）,贡献率分别为12%和6%,来自元江普野的等位基因能提高群体的耐热性,分别可增加9.13%和6.71%的结实率。而位于第8、10染色体上的2个QTL（qHT8和qHT10）,贡献率均为6%,来自元江普野的等位基因降低群体的耐热性,加性效应分别为-6.44%和-4.44%。研究结果不仅为耐热水稻品种的分子标记辅助育种提供参考,而且充分显示了利用野生稻的优异基因改良栽培稻抗逆性状的巨大潜力。     

Populations of methanogenic bacteria in paddy field soil under double cropping conditions (rice-wheat)

The methanogenic populations able to use H₂–CO₂, methanol, and acetate were investigated in paddy field soil in situ under double cropping conditions [rice (Oryza sativa L.) as a summer crop under flooded conditions and wheat (Triticum aestivum L.) as an upland winter crop] over 2 years approximately bimonthly by the most probable number method. Three fields, one without fertilizer, one treated with inorganic fertilizer (mixed fertilizer including urea, ammonium phosphate, and potassium sulfate), and one treated with wheat straw plus inorganic fertilizer, were examined. The population of H₂–CO₂, methanol, and acetate utilizers in the paddy field soil at a depth of 1–6 cm was 10³–10⁴, 10⁴–10⁵, and 10⁴–10⁵ g^-1 dry soil, respectively. These values were almost constant during the 2 years irrespective of moisture regime (flooded or nonflooded), crop (rice or wheat), fertilizer treatment, and soil depth (0–1, 1–10, and 10–20 cm).     

Nitrogen balance in a paddy field planted with whole crop rice (Oryza sativa cv. Kusahonami) during two rice-growing seasons

This paper focuses on N balance in a paddy field planted with whole crop rice (Oryza sativa cv. Kusahonami). The experiment was conducted with two treatments during two rice-growing seasons: one was fertilized with N (160 kg N ha^–1; 16N plot) and the other unfertilized (0N plot); both plots were fertilized with P and K. The N input from precipitation was 15 and 12 kg N ha^–1 in 2002 and 2003, respectively. The N input from irrigation water reached as much as 123 and 69 kg N ha^–1 in 2002 and 2003, respectively. This was because irrigation water contained higher NO₃^– concentrations ranging from 4 to 8 mg N l^–1. The N uptake by rice plants was the major output: 118 and 240 kg N ha^–1 in the 0N and 16N plots in 2002 and 103 and 238 kg N ha^–1 in 2003, respectively. N losses by leaching were 4.8–5.3 and 6.5–7.3 kg N ha^–1 in 2002 and in 2003, respectively. Laboratory experiments were carried out to estimate the amounts of N₂ fixation and denitrification. Amount of N₂ fixation was 43 and 0 kg N ha^–1 in the 0N and 16N plots, respectively. Denitrification potential was quite high in both the plots, and 90% of the N input through irrigation water was lost through denitrification. Collectively, the total N inputs were relatively large due to irrigation water contaminated with NO₃^–, but N outflow loading, expressed as leaching–(irrigation water + precipitation + fertilizer), showed large negative values, suggesting that the whole crop rice field might serve as a constructed wetland for decreasing N.     

Defining the Overall Quality of Cowpea‐Enriched Rice‐Based Breakfast Cereals

The development of innovative legume‐enriched rice products is a promising way to exploit rice varieties with a low sensory grade. In this work, a multidisciplinary approach was applied to the characterization of extruded breakfast cereals prepared from African‐grown Oryza glaberrima (cv. Viwonor) or Oryza sativa (cv. Jasmine 85) enriched with 30% cowpea flour, obtained from sprouted or nonsprouted cowpea. Regardless of the rice species, addition of sprouted cowpea flour conferred a peculiar volatiles profile, rich in sour, bitter, and astringent taste. Protein structural indices provided molecular insights about the macroscopic differences among samples. Extruded products from O. glaberrima were characterized by lower expansion rates with respect to those obtained from O. sativa , regardless of the type of cowpea flour. Sprouting time had a positive influence on the hardness of extruded glaberrima‐based products, facilitating formation of a more compact matrix, but it did not influence sativa‐ based products. Therefore, the breakdown of protein during sprouting appeared fundamental for the incorporation of legume proteins in more compact matrices, such as the one from sativa rice. In the glaberrima‐based products, addition of sprouted cowpea resulted in further loosening of the structure, and this was more evident at increased sprouting times.     

Effects of crop growth and soil treatments on microbial C,N, and P in dry tropical arable land

We studied the dynamics of microbial C, N, and P in soil cropped with rice (Oryza sativa) and lentils (Lens culinaris) in a dryland farming system. The crop biomass and grain yield were also studied. The microbial biomass and its N and P contents were larger under the lentil than under the rice crop. Microbial nutrients decreased as the crops grew and then increased again. Farmyard manure and NPK fertilizer applications increased the level of microbial nutrients, crop biomass, and grain yield by 35–80%, 55–85%, and 74–86%, respectively. However, these applications had no significant effect on most of the soil physicochemical properties in the short term. The microbial biomass was correlated with the crop biomass and grain yield. The calculated flux of N and P through the microbial biomass ranged from 30–45 and 10–19 kg ha^-1 year^-1, respectively. Cultivation of a cereal crop followed by a leguminous crop sustains higher levels of microbial nutrients and hence greater fertility in impoverished tropical arable soils. The soil microbial biomass appears to contribute significantly to crop productivity by releasing nutrients, and applications of manure, either alone or with fertilizers, promote this effect more strongly than the application of NPK fertilizers alone.