Japonica Cv Research Articles

Aluminum triggers broad changes in microRNA expression in rice roots

MicroRNAs are small 21-nucleotide RNA molecules with regulatory roles in development and in response to stress. Expression of some plant miRNAs has been specifically associated with responses to abiotic stresses caused by cold, light, iron, and copper ions. In acid soils, aluminum solubility increases, thereby causing severe damage to plants. Although physiological aspects of aluminum toxicity in plants have been well characterized, the molecular mediators are not fully elucidated. There have been no reports about miRNA responses to aluminum stress. Modulation of miRNA expression may constitute a key element to explain the mechanisms implicated in aluminum toxicity and tolerance. We examined the expression of at least one miRNA member from each miRNA family in rice roots of Oryza sativa spp indica cv. Embrapa Taim and Oryza sativa spp japonica cv. Nipponbare under high concentrations of aluminum. Forty-six miRNA families were effectively detected by quantitative PCR. Among these, 13 were down-regulated and six were up-regulated in roots of the Nipponbare cultivar after 8 h of aluminum treatment. In roots of the Embrapa Taim cultivar, five miRNAs were down-regulated and three were up-regulated. Analyses of their putative targets suggest that these rice miRNAs are involved in the regulation of various metabolic pathways in response to high concentrations of aluminum.

Identification of Japonica Chromosome Segments Associated with Heterosis for Yield in Indica × Japonica Rice Hybrids

To exploit the heterosis for yield and yield‐related traits expressed in indica × japonica F1 hybrids of rice (Oryza sativa L.), a set of 64 chromosome segment substitution lines (CSSLs) was constructed, with each line selected to carry one or a few chromosome segments of the japonica cv. C418 in a genetic background of indica cv. 9311. The CSSLs were crossed one by one with the recipient parent to generate a set of 64 corresponding F1 hybrids (CSSLHs). These materials were field tested over 2 yr for six yield and yield‐related traits. A total of 48 (in 2008) and 55 (in 2009) quantitative trait loci (QTL) were detected, with most individually explaining only a modest proportion of the phenotypic variance. In 2008, 72.9% of the QTL were overdominant, whereas in 2009, this proportion was only 54.5%, indicating that overdominance is important in determining heterosis in japonica × indica rice hybrids, and chromosome segments substitute technology is effective in the utilization of heterosis from intersubspecific cross between indica and japonica rice. Moreover, some of the CSSLs harbored QTL associated with heterosis in both years; these should represent potential candidates as parents of F1 hybrids along with cv. 9311 or other indica lines.

Expression Analysis of miRNAs and Highly‐expressed Small RNAs in Two Rice Subspecies and their Reciprocal Hybrids

Heterosis, or hybrid vigor, is the phenomenon whereby progeny of two inbred lines exhibit superior agronomic performance compared with either parent. We analyzed the expression of miRNAs and highly expressed small RNAs (defined according to Solexa sequencing results) in two rice (Oryza sativa) subspecies (japonica cv. Nipponbare and indica cv. 93-11) and their reciprocal hybrids using microarrays. We found that of all the 1141 small RNAs tested, 140 (12%, 140 of 1141) and 157 (13%, 157 of 1141) were identified being significantly differentially expressed in two reciprocal hybrids, respectively. All possible modes of action, including additive, high- and low- parent, above high- and below low-parent modes were exhibited. Both F1 hybrids showed non-additive expression patterns, with downregulation predominating. Interestingly, 15 miRNAs displayed stark opposite expression trends relative to mid-parent in reciprocal hybrids. Computational prediction of targets of differentially expressed miRNAs showed that they participated in multifaceted developmental pathways, and were not distinguishable from the targets of non-differentially expressed miRNAs. Together, our findings reveal that small RNAs play roles in heterosis and add a new layer in the understanding and exploitation of molecular mechanisms of heterosis.

Marker-assisted breeding of a photoperiod-sensitive male sterile japonica rice with high cross-compatibility with indica rice

The incomplete fertility of japonica × indica rice hybrids has inhibited breeders’ access to the substantial heterotic potential of these hybrids. As hybrid sterility is caused by an allelic interaction at a small number of loci, it is possible to overcome it by simple introgression at the major sterility loci. Here we report the use of marker-assisted backcrossing to transfer into the elite japonica cv. Zhendao88 a photoperiod-sensitive male sterility gene from cv. Lunhui422S (indica) and the yellow leaf gene from line Yellow249 (indica). The microsatellite markers RM276, RM455, RM141 and RM185 were used to tag the fertility genes S5, S8, S7 and S9, respectively. Line 509S is a true-breeding photoperiod-sensitive male sterile plant, which morphologically closely resembles the japonica type. Genotypic analysis showed that the genome of line 509S comprises about 92% japonica DNA. Nevertheless, hybrids between line 509S and japonica varieties suffer from a level of hybrid sterility, although the line is highly cross-compatible with indica types, with the resulting hybrids expressing a significant degree of heterosis. Together, these results suggest that segment substitution on fertility loci based on known information and marker-assisted selection are an effective approach for utilizing the heterosis of rice inter-subspecies.

Fine mapping of grain weight QTLs using near isogenic lines from a Cross between Oryza sativa and O. grandiglumis

In our previous study, we reported the grain weight (GW) QTL, tgw11 in isogenic lines derived from a cross between Oryza sativa ssp. Japonica cv. Hwaseong and O. grandiglumis. The O. grandiglumis allele at tgw11 decreased GW in the Hwaseong background. To fine-map tgw11, one F5 plant homozygous for the O. grandiglumis DNA in the target region on chromosome 11 was selected from F4 line, CR1242 segregating for tgw11 and crossed with Hwaseong to produce secondary F2 and F3 populations. QTL analysis using 760 F2 plants confirmed the existence of tgw11 with an R2 value of 15.0%. This QTL explained 32.2% of the phenotypic variance for GW in 91 F3 lines. Substitution mapping with 65 F3 lines with informative recombination breakpoints in the target region was carried out to narrow down the position of the tgw11. The result indicated that tgw11 was located in the 900-kb interval between two SSR markers, RM224 and RM27358. QTLs for grain width and grain thickness were also located in the same interval suggesting that a single gene is involved in controlling these three traits. Analysis of F3 lines indicated that the variation in TGW is associated with variation in grain shape, specifically grain thickness and grain width. Genetic analysis indicated that the O. grandiglumis allele for small seed was dominant over the Hwaseong allele. SSR markers tightly linked to the GW QTL would be useful in marker-assisted selection for variation in GW in breeding program.

QTL analysis for flowering time using backcross population between Oryza sativa Nipponbare and O. rufipogon

In the near future, global average temperature is expected to increase due to the accumulation of greenhouse gases, and increased temperatures will cause severe sterility in many crop species. In rice, since wild species show high genetic variation, they may have the potential to improve the flowering characters of cultivars. In this study, we investigated flowering characters under natural conditions by comparing an Asian wild rice accession of Oryza rufipogon W630 (originated from Myanmar) with a Japanese rice cultivar, O. sativa Japonica cv. Nipponbare. Further, QTL analysis for days to heading (DH) and spikelet opening time (SOT: the time of day when the spikelet opens) was carried out using BC(2)F(8) backcross population derived from the cross between them. Regarding DH, four QTLs were detected, and two of them were found to have wild alleles with strong effects leading to longer days to heading during the Japanese summer. These wild alleles may be used to produce late-heading cultivars that do not flower during the high summer temperatures anticipated in the future. As for SOT, two parameters of SOTb (beginning time when the first spikelet opens) and SOTm (median time when 50% of the spikelets open) were recorded and the time differences from Nipponbare were investigated. Two QTLs on chromosomes 5 and 10 and two QTLs on chromosomes 4 and 5 were detected for SOTb and SOTm, respectively. The wild alleles were responsible for early spikelet opening time at all loci. If the wild alleles detected in this study have the same effects in the genetic background of other cultivars, they will be very useful in producing early-flowering rice cultivars that complete fertilization in the morning before the temperature rises.

Systematic identification of X1-homologous genes reveals a family involved in stress responses in rice

X1-homologous genes (XHS) encode plant-specific proteins containing three major domains (XH, XS, zf-XS), but their functions are largely unknown. We report the systematic identification and characterization of XHS genes in the rice genome. Eleven putative XHS protein sequences (OXHS1-11) were identified in the sequenced genome of Oryza sativa japonica cv. Nipponbare, and these sequences, along with other plant XHS homologues, were classified into five subgroups based on phylogenetic analysis. Distinct diversification of the XHS proteins occurred between monocotyledon and dicotyledon plants. The OXHS family has diverse exon-intron structures and organizations of putative domains and motifs. The OXHS proteins showed no transactivation activity, and no interaction between the XH domain and the XS domain in yeast. Four representative OXHS proteins were targeted to cytoplasm, which contradicts the previous speculation that XHS proteins are putative transcription factors. All the OXHS genes are predominantly expressed in floral organs, and some are expressed in a wide range of tissues or organs in indica rice Minghui 63. Nine OXHS genes are responsive to at least one of the abiotic stresses including drought, salt, cold, and abscisic acid treatment. Over-expression of one stress-responsive gene OXHS2 in rice resulted in reduced tolerance to salt and drought stresses. These results suggest that the OXHS family may be functionally diversified and some members of this family may play important roles in regulating stress tolerance in rice.

Fine genetic mapping and physical delimitation of the lesion mimic gene spotted leaf 5 (spl5) in rice (Oryza sativa L.)

Spotted leaf 5 (spl5), a lesion mimic mutant, was first identified in rice (Oryza sativa L.) japonica cv. Norin8 in 1978. This mutant exhibits spontaneous disease-like lesions in the absence of any pathogens and resistance to rice blast and bacterial blight; however, the target gene has not yet been isolated. In the present study, we employed a map-based cloning strategy to finely map the spl5 gene. In an initial mapping with 100 F2 individuals (spl5/spl5) derived from a cross between the spl5 mutant and indica cv. 93-11, the spl5 gene was located in a 3.3-cM region on chromosome 7 using six simple sequence repeat (SSR) markers. In a high-resolution genetic mapping, two F2 populations with 3,149 individuals (spl5/spl5) were derived from two crosses between spl5 mutant and two indica cvs. 93-11 and Zhefu802 and six sequence-tagged site (STS) markers were newly developed. Finally, the spl5 gene was mapped to a region of 0.048 cM between two markers SSR7 and RM7121. One BAC/PAC contig map covering these markers’ loci and the spl5 gene was constructed through Pairwise BLAST analysis. Our bioinformatics analysis shows that the spl5 gene is located in the 80-kb region between two markers SSR7 and RM7121 with a high average ratio of physical to genetic distance (1.67 Mb/cM) and eighteen candidate genes. The analysis of these candidate genes indicates that the spl5 gene represents a novel class of regulators controlling cell death and resistance response in plants.

Determination of Resistance Spectra of the Pi-ta and Pi-k Genes to U.S. Races of Magnaporthe oryzae Causing Rice Blast in a Recombinant Inbred Line Population.

Molecular tagged resistance (R) genes are useful for developing improved cultivar resistance using marker-assisted breeding. In the present study, R genes to common races of Magnaporthe oryzae, the causal agent of blast disease of rice (Oryza sativa), were mapped using an F10 recombinant inbred line (RIL) population derived from a cross of tropical japonica cv. Katy with breeding line RU9101001. Katy was resistant to 10 common U.S. races: IA-45, IB-1, IB-45, IB-49, IB-54, IC-17, ID-1, IE-1, IG-1, and IH-1 of M. oryzae. RU9101001 was resistant to races IA-45, IB-45, IB-54, IG-1, and IH-1. Katy and RU9101001 were susceptible to race IE-1k. Twenty-three polymorphic simple sequence repeat (SSR) markers were used to map R genes. Segregation ratios of 1:1 (resistant/susceptible) to races IB-1, IB-49, IC-17, ID-1, and IE-1 indicated the presence of a single dominant R gene in Katy. Ratios of 3:1 (resistant/susceptible) to races IA-45, IB-45, IG-1, and IH-1 indicated that a single R gene was present in Katy and a different R gene was present in RU9101001. Resistance to the abovementioned races was correlated with the presences of the Pi-ta gene and 11 Katy SSR alleles, suggesting that Pi-ta confers resistance to IA-45, IB-1, IB-45, IB-49, IC-17, IG-1, ID-1, IE-1, and IH-1. Katy, RU9101001, and all RILs were resistant to race IB-54, which was consistent with the presence of Pi-ks in Katy and Pi-kh in RU9101001. Resistance to IA-45, IB-45, IG-1, and IH-1 correlated with the presence of Pi-kh, suggesting that Pi-kh confers resistance to IA-45, IB-45, IG-1, and IH-1. These data suggest that Pi-ta and Pi-kh are effective R genes with overlapped resistance to the 10 common races of M. oryzae.

Quantitative Trait Loci Analysis of Responses to Nitrogen and Phosphorus Deficiency in Rice Chromosomal Segment Substitution Lines

Tolerance to low nitrogen and low phosphorus conditions is a highly desired characteristic for sustainable crop production. In this study, a set of 125 chromosome segment substitution lines (CSSL), each containing a single or few introgression segments from a japonica cv. Nipponbare with the genetic background of an indica cv. 9311, were evaluated using augmented design under the field experiment with normal, low nitrogen (N0), and low phosphorus (P0) conditions. The grain yield and panicle number per plant were measured for each CSSL, and their relative values based on N0 or P0 and normal conditions were considered as the measurement for tolerance to low soil nutrient. The results showed that both the N0 and P0 conditions had strong negative effect on yield and panicle number, and there were different responses among the CSSLs to the stresses, and the relative traits had a significant nega- tive correlation with the traits under the normal condition. 9311 showed better tolerance to low nutrient conditions than Nipponbare. A total of 38 chromosomal regions or quantitative trait loci (QTL) all with negative allelic effects from Nipponbare were detected for the measured traits under the nitrogen and the phosphorus stresses, of those 26 QTLs were for the yield and panicle number, 12 QTLs for the relative traits. Five chromosomal regions were identified in common under both the stresses, while most QTLs (81%) were specifically detected only in low nitrogen or phosphorus condition. Such different QTLs suggest that the responses to limiting nitrogen and phosphorus conditions are regulated by different sets of genes in rice. Most QTLs for the relative traits were co-localized with those for the yield and the panicle number under either nitrogen or phosphorus stresses, indicating that the tolerance QTLs may be involved in nitrogen and phosphorus uptake or assimilation pathway in rice.

Mapping quantitative trait loci for awnness and yield component traits in isogenic lines derived from an Oryza sativa / O. rufipogon cross

An advanced backcross line, HR9118, was produced from a single plant of BC2F3 families derived from a cross between Oryza rufipogon Griff. (IRGC 105491) as a donor parent and the O. sativa subsp. japonica cv. Hwaseongbyeo as a recurrent parent. Although HR9118 resembled Hwaseongbyeo, several traits were different from those of Hwasoengbyeo, including days to heading, plant height, and awn. These differences between Hwasongbyeo and HR9118 could be attributed to introgressed O. rufipogon chromosome segments into HR9118. Introgression analysis using 460 SSR markers revealed that three O. rufipogon-specific chromosome segments were detected in HR9118 genome. F2:3 populations derived from the cross between Hwaseongbyo and HR9118, consisting of 340 F2 plants and 137 F3 lines, were used to map and characterize QTLs for 12 traits. QTL analysis identified a total of 17 QTLs in the F2:3 populations. Of these, seven QTLs were shared by the F2 and F3 populations, whereas the other ten QTLs were identified only in the F3 population. In seven (41.2%) QTLs identified in this study, the O. rufipogon-derived alleles contributed desirable agronomic effects despite the overall undesirable characteristics of the wild phenotype. Each of three O. rufipogon introgressed segments contained multiple QTLs, indicating linkage and/or pleotropic effects. A cluster of eight QTLs was detected on chromosome 8 including a major QTL for awn. Substitution mapping using F2 population indicated that awn8 was located within an interval between two SSR makers RM23326 and RM23356 which are 590 kb apart. SSR markers tightly linked to QTLs for yield components detected in this study will facilitate cloning of the gene underlying this QTL as well as marker-assisted selection for variation in grain weight in an applied breeding program.

Occurrence of D-serine in rice and characterization of rice serine racemase

Germinated, unpolished rice was found to contain a substantial amount of D-serine, with the ratio of the D-enantiomer to the L-enantiomer being higher for serine than for other amino acids. The relative amount of D-serine (D/(D + L)%) reached approximately 10% six days after germination. A putative serine racemase gene ( serr, clone No. 001-110-B03) was found in chromosome 4 of the genomic DNA of Oryza sativa L. ssp. Japonica cv. Nipponbare. This was expressed as serr in Escherichia coli and its gene product (SerR) was purified to apparent homogeneity. SerR is a homodimer with a subunit molecular mass of 34.5 kDa, and is highly specific for serine. In addition to a serine racemase reaction, SerR catalyzes D- and L-serine dehydratase reactions, for which the specific activities were determined to be 2.73 and 1.42 nkatal/mg, respectively. The optimum temperature and pH were respectively determined for the racemase reaction (35 °C and pH 9.0) and for the dehydratase reaction (35 °C and pH 9.5). SerR was inhibited by PLP-enzyme inhibitors. ATP decreased the serine racemase activity of SerR but increased the serine dehydratase activity. Kinetic analysis showed that Mg 2+ increases the catalytic efficiency of the serine racemase activity of SerR and decreases that of the serine dehydratase activity. Fluorescence-quenching analysis of the tryptophan residues in SerR indicated that the structure of SerR is distorted by the addition of Mg 2+, and this structural change probably regulates the two enzymatic activities.

Molecular cloning and characterization of OsCDase, a ceramidase enzyme from rice.

Sphingolipids are a structurally diverse group of molecules based on long-chain sphingoid bases that are found in animal, fungal and plant cells. In contrast to the situation in animals and yeast, much less is known about the spectrum of sphingolipid species in plants and the roles they play in mediating cellular processes. Here, we report the cloning and characterization of a plant ceramidase from rice (Oryza sativa spp. Japonica cv. Nipponbare). Sequence analysis suggests that the rice ceramidase (OsCDase) is similar to mammalian neutral ceramidases. We demonstrate that OsCDase is a bona fide ceramidase by heterologous expression in the yeast double knockout mutant Deltaypc1Deltaydc1 that lacks the yeast ceramidases YPC1p and YDC1p. Biochemical characterization of OsCDase showed that it exhibited classical Michaelis-Menten kinetics, with optimum activity between pH 5.7 and 6.0. OsCDase activity was enhanced in the presence of Ca(2+), Mg(2+), Mn(2+) and Zn(2+), but inhibited in the presence of Fe(2+). OsCDase appears to use ceramide instead of phytoceramide as a substrate. Subcellular localization showed that OsCDase is localized to the endoplasmic reticulum and Golgi, suggesting that these organelles are sites of ceramide metabolism in plants.

Ammonium under solution culture alleviates aluminum toxicity in rice and reduces aluminum accumulation in roots compared with nitrate

Al stress and ammonium–nitrogen nutrition often coexist in acidic soils due to their low pH and weak nitrification ability. Rice is the most Al-resistant species among small grain cereal crops and prefers NH4+ as its major inorganic nitrogen source. This study investigates the effects of NH4+ and NO3− on Al toxicity and Al accumulation in rice, and thereby associates rice Al resistance with its NH4+ preference. Two rice subspecies, indica cv. Yangdao6 and japonica cv. Wuyunjing7, were used in this study. After treatment with or without Al under conditions of varying NH4+ and NO3− supply, rice seedlings were harvested for the determination of root elongation, callose content, biomass, Al concentration and medium pH. The results indicated that Wuyunjing7 was more Al-resistant and NH4+-preferring than Yangdao6. NH4+ alleviated Al toxicity in two cultivars compared with NO3−. Both NH4+-Al supply and pretreatment with NH4+ reduced Al accumulation in roots and root tips compared with NO3−. NH4+ decreased but NO3− increased the medium pH, and root tips accumulated more Al with a pH increase from 3.5 to 5.5. Increasing the NO3− concentration enhanced Al accumulation in root tips but increasing the NH4+ concentration had the opposite effect. These results show NH4+ alleviates Al toxicity for rice and reduces Al accumulation in roots compared with NO3−, possibly through medium pH changes and ionic competitive effects. Making use of the protective effect of NH4+, in which the Al resistance increases, is advised for acidic soils, and the hypothesis that rice Al resistance is associated with the preferred utilization of NH4+ is suggested.

Gene Structure and Expression of the High‐affinity Nitrate Transport System in Rice Roots

Rice has a preference for uptake of ammonium over nitrate and can use ammonium-N efficiently. Consequently, transporters mediating ammonium uptake have been extensively studied, but nitrate transporters have been largely ignored. Recently, some reports have shown that rice also has high capacity to acquire nitrate from growth medium, so understanding the nitrate transport system in rice roots is very important for improving N use efficiency in rice. The present study identified four putative NRT2 and two putative NAR2 genes that encode components of the high-affinity nitrate transport system (HATS) in the rice (Oryza sativa L. subsp. japonica cv. Nipponbare) genome. OsNRT2.1 and OsNRT2.2 share an identical coding region sequence, and their deduced proteins are closely related to those from mono-cotyledonous plants. The two NAR2 proteins are closely related to those from mono-cotyledonous plants as well. However, OsNRT2.3 and OsNRT2.4 are more closely related to Arabidopsis NRT2 proteins. Relative quantitative reverse transcription-polymerase chain reaction analysis showed that all of the six genes were rapidly upregulated and then downregulated in the roots of N-starved rice plants after they were re-supplied with 0.2 mM nitrate, but the response to nitrate differed among gene members. The results from phylogenetic tree, gene structure and expression analysis implied the divergent roles for the individual members of the rice NRT2 and NAR2 families. High-affinity nitrate influx rates associated with nitrate induction in rice roots were investigated and were found to be regulated by external pH. Compared with the nitrate influx rates at pH 6.5, alkaline pH (pH 8.0) inhibited nitrate influx, and acidic pH (pH 5.0) enhanced the nitrate influx in 1 h nitrate induced roots, but did not significantly affect that in 4 to 8 h nitrate induced roots.

Introgression of a quantitative trait locus for spikelets per panicle from Oryza minuta to the O. sativa cultivar Hwaseongbyeo

AbstractA new QTL for spikelets per panicle (SPP) was detected on the long arm of chromosome 7 in an F2 population derived from a cross between the japonica cultivar Hwaseongbyeo and WH29001. WH29001, an advanced backcross line was developed by introgressing chromosomal segments from an accession of O. minuta (2n = 48, BBCC, Acc. no. 101141) into the O. sativa subsp. japonica cv. Hwaseongbyeo. The O. minuta allele increased SPP in the Hwaseongbyeo background despite the fact that O. minuta was the small panicle parent. Using F3 and F4 progenies, spp7 was validated and mapped to a 2.3 Mb region in the interval between the SSR markers RM445 and RM21615 based on the japonica genome sequence. A yield trial using F4 lines indicated that the lines carrying an O. minuta chromosome segment across the entire spp7 target region out‐yielded its sister lines containing Hwaseongbyeo chromosome in the target region and Hwaseongbyeo by 14.3% and 15.9%, respectively. Increase in SPP in WH29001 was mainly because of the increase in primary branches per panicle. The locus, spp7 is of particular interest because of its independence from undesirable height and flowering time. SSR markers tightly linked to the spp7 will facilitate cloning of the gene underlying this QTL as well as marker‐assisted selection for variation in SPP in an applied breeding program.

Resistance to Rice yellow mottle virus in rice germplasm in Madagascar

Rice yellow mottle virus (RYMV) is a recent and major threat to rice production in Madagascar. A large scale screening of resistance to RYMV in rice germplasm in Madagascar was conducted by visual symptom scoring and virus-assessment through ELISA tests. The response to virus inoculation of 503 local or introduced rice accessions was assessed. Most of them were susceptible to RYMV. A few cultivars expressed partial resistance at a level similar to the partially resistant Oryza sativa japonica cv. Azucena. Only one O. sativa cultivar expressed high resistance characterised by a lack of symptoms and an undetectable level of virus. It was a Malagasy traditional indica cultivar, named Bekarosaka, which originated from the northwest of the country. It was selected by farmers for its field resistance to RYMV. The response of cv. Bekarosaka to four pathotypes of RYMV was similar to that of cv. Gigante, the only other highly resistant indica cultivar. The sequence of the middle domain of the eIF(iso)4G, the genetic determinant of Rymv1 resistance on chromosome 4, of cv. Bekarosaka was similar to that of cv. Gigante. Subsequently, cvs Bekarosaka and Gigante probably carried the same resistance allele Rymv1-2. Rymv1-2 resistance was efficient against isolates of the major strains of RYMV, but was readily overcome by a pathotype from the northwest of Madagascar.

Relationships between Phosphatidylglycerol Molecular Species of Thylakoid Membrane Lipids and Sensitivities to Chilling‐induced Photoinhibition in Rice

In an attempt to explore the relationships between phosphatidylglycerol (PG) molecular species of thylakoid membrane lipids and sensitivities to chilling-induced photoinhibition, PG molecular species, D1 protein, electron transport activities of thylakoid membrane and the potential quantum yield (F(v)/F(m)) in rice treated under middle and low photon flux density (PFD) at 11 degrees C were analyzed by high performance liquid chromatography, enzyme hydrolysis, gas phase chromatography (GC) and so on. Results showed that the major molecular species of PGs in rice thylakoid membrane were 18:3/16:0, 18:3/16:1(3t), 18:2/16:0, 18:2/16:1(3t), 18:1/16:0, 18:1/16:1(3t), 16:0/16:0, 16:0/16:1(3t). There were large differences in the contents of unsaturated PG molecular species such as 18:1 approximately 3/16:0 approximately 16:1(3t) and saturated PG molecular species like 16:0/16:0 approximately 16:1(3t) among japonica cv 9516 (j-9516), japonica-indica hybrid F1 j-9516/i-SY63 (ji-95SY) and indica cv Shanyou 63 (i-SY63). J-9516 containing higher contents of unsaturated PG molecular species was manifest in stable D1 protein contents under chill and tolerant to chill-induced photoinhibition. In contrast to j-9516, i-SY63 with lower contents of unsaturated PG molecular species, exhibited unstable D1 protein contents under chill and was sensitive to chill-induced photoinhibition. ji-95SY containing middle contents of unsaturated PG molecular species between those of j-9516 and i-SY63, exhibited mid extent of sensitivity to chill-induced photoinhibition. The losses in D1 protein also account for the inhibition in electron transport activity of thylakoid membrane and the observed decline in F(v)/F(m). The PG molecular species that is efficient in raising chilling-resistant capacity were those containing unsaturated fatty acids, namely, unsaturated PG molecular species. These results implied that the substrate selectivity of the glycerol-3-phosphate acyltransferase in chloroplasts towards 16:0 or 18:1 displayed greatly the difference between japonica and indica rice. It was possible to enhance the capacity of resistance to chilling-induced photoinhibition by improving or modifying the GPAT gene.

Transient Expression of Green Fluorescent Protein in Rice Calluses : Optimization of Parameters for Helios Gene Gun Device

An optimized condition for particle bombardment is necessary for efficient genetic transformation. Parameters for Helios gene gun, the new system for nucleic acid delivery which is mainly consists of hand-held device sold by Bio-Rad Laboratories (California USA), were examined based on transient expression of synthetic green fluorescent protein (sgfp) in rice calluses of indica cv. Fatmawati and japonica cv. Nipponbare. In the experimental conditions that we examined, parameters found to be the most favorable conditions for transient expression of sgfp in rice callus cells were as follows: 200−250 psi helium pressure, 0.6 μm gold particle size, 0.25 mg gold particles per shot, and 1.5 μg plasmid-DNA per shot. Desiccation of callus cells for eight min was also found appropriate. The level of transient sgfp expression was not significantly influenced by the pre-culture for 4 to 12 d before bombardment or by callus age between 10 and 33 wk old in Fatmawati. These parameters for this particular device should improve the transient expression, thus enabling stable expression of introduced genes via Helios gene gun using callus as a target tissue.

The hybrid breakdown 1(t) locus induces interspecific hybrid breakdown between rice Oryza sativa cv. Koshihikari and its wild relative O. nivara

Reproductive barriers are often observed in progenies derived from crosses between relatively distant species, such as cultivated varieties and their wild relatives. Such barriers hinder the transfer of agronomically important genes from one species to another. In the process of constructing a series of introgression lines (ILs) between Oryza sativa japonica cv. Koshihikari and wild species O. nivara, we observed the occurrence of hybrid breakdown, a reproductive barrier observed in F2 and later generations. Plants affected by this hybrid breakdown show extremely weak growth and die before heading. Linkage analysis and construction of ILs indicated that the locus -hbd1(t), located on the short arm of chromosome 2 of O. nivara, induces hybrid breakdown in the Koshihikari genetic background. hbd1(t) is identical to hbd1, which was previously identified in a cross between Koshihikari and Nona Bokra (indica) cultivars of O. sativa. We present genetic evidence that hbd1(t) regulates hybrid breakdown and its location within 50 kb. Possible reasons for this locus being responsible for hybrid breakdown in inter- and intraspecific crosses are discussed.