Mature Rice Seeds Research Articles

Biofortified Rice Provides Rich Sakuranetin in Endosperm

Sakuranetin plays a key role as a phytoalexin in plant resistance to biotic and abiotic stresses, and possesses diverse health-promoting benefits. However, mature rice seeds do not contain detectable levels of sakuranetin. In the present study, a transgenic rice plant was developed in which the promoter of an endosperm-specific glutelin gene OsGluD-1 drives the expression of a specific enzyme naringenin 7-O-methyltransferase (NOMT) for sakuranetin biosynthesis. The presence of naringenin, which serves as the biosynthetic precursor of sakuranetin made this modification feasible in theory. Liquid chromatography tandem mass spectrometry (LC–MS/MS) validated that the seeds of transgenic rice accumulated remarkable sakuranetin at the mature stage, and higher at the filling stage. In addition, the panicle blast resistance of transgenic rice was significantly higher than that of the wild type. Specially, the matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) imaging was performed to detect the content and spatial distribution of sakuranetin and other nutritional metabolites in transgenic rice seeds. Notably, this genetic modification also did not change the nutritional and quality indicators such as soluble sugars, total amino acids, total flavonoids, amylose, total protein, and free amino acid content in rice. Meanwhile, the phenotypes of the transgenic plant during the whole growth and developmental periods and agricultural traits such as grain width, grain length, and 1000-grain weight exhibited no significant differences from the wild type. Collectively, the study provides a conceptual advance on cultivating sakuranetin-rich biofortified rice by metabolic engineering. This new breeding idea may not only enhance the disease resistance of cereal crop seeds but also improve the nutritional value of grains for human health benefits.

Transcriptomic Analysis of the Dehydration Rate of Mature Rice (Oryza sativa) Seeds.

In this study, a transcriptomic analysis of the dehydration rate of mature rice seeds was conducted to explore candidate genes related to the dehydration rate and provide a theoretical basis for breeding and utilization. We selected two rice cultivars for testing (Baghlani Nangarhar, an extremely rapid dehydration genotype, and Saturn, a slow dehydration genotype) based on the results determined by previous studies conducted on the screening of 165 germplasm materials for dehydration rate phenotypes. A rapid dehydration experiment performed on these two types of seeds was conducted. Four comparative groups were set up under control and dehydration conditions. The differentially expressed genes (DEGs) were quantified via transcriptome sequencing and real-time quantitative PCR (RT-qPCR). GO (Gene ontology) and KEGG(Kyoto Encyclopedia of Genes and Genomes) analyses were also conducted. In Baghlani Nangarhar, 53 DEGs were screened, of which 33 were up-regulated and 20 were down-regulated. In Saturn, 25 DEGs were screened, of which 19 were up-regulated and 6 were down-regulated. The results of the GO analysis show that the sites of action of the differentially expressed genes enriched in the rapid dehydration modes are concentrated in the cytoplasm, internal components of the membrane, and nucleosomes. They play regulatory roles in the processes of catalysis, binding, translocation, transcription, protein folding, degradation, and replication. They are also involved in adaptive responses to adverse external environments, such as reactive oxygen species and high temperature. The KEGG analysis showed that protein processing in the endoplasmic reticulum, amino acid biosynthesis, and oxidative phosphorylation were the main metabolic pathways that were enriched. The key differentially expressed genes and the most important metabolic pathways identified in the rapidly and slowly dehydrated genotypes were protein processing in the endoplasmic reticulum and oxidative phosphorylation metabolism. They were presumed to have important regulatory roles in the mechanisms of stress/defense, energy metabolism, protein synthesis/folding, and signal transduction during the dehydration and drying of mature seeds. The results of this study can potentially provide valuable information for further research on the genes and metabolic pathways related to the dehydration rate of mature rice seeds, and provide theoretical guidance for the selection and breeding of new rice germplasm that can be rapidly dehydrated at the mature stage.

Efeitos de fatores ambientais na germinação sazonal de diferentes biótipos de arroz vermelho (Oryza sativa L.)

ABSTRACT: Weedy biotypes of Oryza sativa L., (weedy rice) are a serious threat to rice production because of their flexibility in seed germination timing, variable growth forms, and high genetic diversity. Experiments were designed to determine the effects of storage conditions, osmotic stress, pH, salt stress, and burial depth on germination of weedy rice seeds in flooded or non-flooded conditions. Mature weedy rice seeds were gathered from rice fields in the different regions of Turkey. Three biotypes were selected and classified according to awn length; long awn, short awn and awnless. Seasonal germination patterns of weedy rice seeds in the laboratory after retrieval from various depths and timing were affected by burial depth, water regime, and exhumation timing across the treatments for all biotypes. The long and short awn biotypes had higher germination rates than the awnless biotype and did not have a seasonal germination pattern. Seed germination was initiated in the spring, peaked in summer and declined in the fall in flooded and non-flooded treatments for the awnless biotype. The most distinct differences in seedling emergence patterns were observed in awnless and long awn biotypes, and they emerged from 10 cm depth. Our results suggested that flooded conditions reduced the germination of weedy rice biotypes. Flooding rice paddies for a period of time after harvest may improve weedy rice control and decrease the weed population. In addition to deeper cultivation would be more effective in controlling all weedy biotypes since the majority of weed seedling emergence was from shallow depths.

Appraisal of genetic variability in sodium azide induced rice mutants to identify selection criteria for higher yield using quantitative attributes

Induced mutation is an effective tool in generating variability of crop plants. Identification of efficient genotypes with improved yield requires knowledge of genetic variation in yield and yield contributing traits. Therefore, an investigation was conducted to develop variants through azide mutagenesis and estimation of genetic variability of the mutants to detect selection standards towards higher yield. Mature rice seeds were mutagenized with estimated LD50 concentration. A hundred mutants were grown-up accordingly and their quantitative traits were evaluated through multivariate analysis to assess genetic variability. Every assessed trait except grain length exhibited highly significant variation for all mutants. The high genotypic and phenotypic coefficient of variation along with high heritability and genetic advance as a % of mean was accounted for total and filled grains panicle-1. Grain yield was positively and significantly correlated with total grains panicle-1, filled grains panicle-1 and straw yield hill-1. Besides, higher variance was also associated with these traits. Ward’s Agglomerative clustering grouped the mutants into 7 major clusters. More than 36% of total variation was associated with first 2 principal components (PCs) and was mostly with total grains panicle-1 and filled grains panicle-1 whereas third and fourth PCs were mainly accounted for straw yield hill-1. Thus, these traits should receive special attention during selection of yield potential genotypes. Presence of genetic variation in mutants was ensured for most of the traits and selection based on greater tillers hill-1, grains panicle-1 and straw yield hill-1 may open a new avenue towards improved yield and other traits in rice.

Callogenesis and somatic embryogenesis of <i>Oryza sativa</i> L. (cv. MARDI Siraj 297) under the influence of 2, 4-dichlorophenoxyacetic acid and kinetin

<abstract> <p>Callogenesis and embryogenesis are integral parts of many tissue culture procedures for genetic manipulation in rice. However, the efficiency of both processes is largely dependent on the media constituent especially the plant growth regulators (PGRs) due to the genotype-dependent nature of <italic>in vitro</italic> culture protocols. Therefore, this study investigates the effect of two PGRs; 2, 4-dichlorophenoxyacetic acid (2, 4-D) and kinetin (Kin) on callus growth and somatic embryogenesis of an important Malaysian rice cultivar (<italic>Oryza sativa</italic> L. cv. MARDI Siraj 297). Mature rice seeds explants were inoculated in Murashige & Skoog (MS) medium supplemented with different combinations of 2, 4-D (0 to 3.5 mg/L) and Kin (0 to 0.5 mg/L) to induce callogenesis. Parameters for callus growth such as fresh weight (FW), callus induction frequency (CIF), embryogenic callus frequency (ECF), regeneration frequency (RF), number of plantlets per callus (PPC), callus texture and callus color were observed after 35 days of inoculation. The results show that the maximum callus growth was achieved in MS medium supplemented with combination of 2.0 mg/L 2, 4-D and 0.2 mg/L Kin, represented by the highest FW (211 mg), CIF (95%), ECF (90%), RF (100%) and PPC (22 plantlets); along with friable callus texture. Low concentration of 2, 4-D (0 to 0.5 mg/L) in the presence or absence of Kin promotes root growth instead of callus, while high concentrations (above 3.0 mg/L) retard the callus formation. The embryogenic calli from this optimized PGRs combination were successfully formed shoots in MS medium supplemented with 2 mg/L BAP and 1 mg/L NAA, followed by rooting in PGRs-free MS medium. This finding provides an efficient protocol for callogenesis and somatic embryogenesis of MARDI Siraj 297, since this is the first published report regarding somatic embryogenesis induction of this cultivar.</p> </abstract>

Transcriptome comparison between pluripotent and non-pluripotent calli derived from mature rice seeds

In vitro plant regeneration involves a two-step practice of callus formation and de novo organogenesis. During callus formation, cellular competence for tissue regeneration is acquired, but it is elusive what molecular processes and genetic factors are involved in establishing cellular pluripotency. To explore the mechanisms underlying pluripotency acquisition during callus formation in monocot plants, we performed a transcriptomic analysis on the pluripotent and non-pluripotent rice calli using RNA-seq. We obtained a dataset of differentially expressed genes (DEGs), which accounts for molecular processes underpinning pluripotency acquisition and maintenance. Core regulators establishing root stem cell niche were implicated in pluripotency acquisition in rice callus, as observed in Arabidopsis. In addition, KEGG analysis showed that photosynthetic process and sugar and amino acid metabolism were substantially suppressed in pluripotent calli, whereas lipid and antioxidant metabolism were overrepresented in up-regulated DEGs. We also constructed a putative coexpression network related to cellular pluripotency in rice and proposed potential candidates conferring pluripotency in rice callus. Overall, our transcriptome-based analysis can be a powerful resource for the elucidation of the molecular mechanisms establishing cellular pluripotency in rice callus.

Transcriptomics of Mature Rice (Oryza Sativa L. Koshihikari) Seed under Hot Conditions by DNA Microarray Analyses

Higher temperature conditions during the final stages of rice seed development (seed filling and maturation) are known to cause damage to both rice yield and rice kernel quality. The western and central parts of Japan especially have seen record high temperatures during the past decade, resulting in the decrease of rice kernel quality. In this study, we looked at the rice harvested from a town in the central Kanto-plains (Japan) in 2010. The daytime temperatures were above the critical limits ranging from 34 to 38 °C at the final stages of seed development and maturity allowing us to investigate high-temperature effects in the actual field condition. Three sets of dry mature rice seeds (commercial), each with specific quality standards, were obtained from Japan Agriculture (JA Zen-Noh) branch in Ami-town of Ibaraki Prefecture in September 2010: grade 1 (top quality, labeled as Y1), grade 2 (medium quality, labeled as Y2), and grade 3 (out-of-grade or low quality, labeled as Y3). The research objective was to examine particular alterations in genome-wide gene expression in grade 2 (Y2) and grade 3 (Y3) seeds compared to grade 1 (Y1). We followed the high-temperature spike using a high-throughput omics-approach DNA microarray (Agilent 4 × 44 K rice oligo DNA chip) in conjunction with MapMan bioinformatics analysis. As expected, rice seed quality analysis revealed low quality in Y3 > Y2 over Y1 in taste, amylose, protein, and fatty acid degree, but not in water content. Differentially expressed gene (DEG) analysis from the transcriptomic profiling data revealed that there are more than one hundred upregulated (124 and 373) and downregulated (106 and 129) genes in Y2 (grade 2 rice seed) and Y3 (grade 3 rice seed), respectively. Bioinformatic analysis of DEGs selected as highly regulated differentially expressed (HRDE) genes revealed changes in function of genes related to metabolism, defense/stress response, fatty acid biosynthesis, and hormones. This research provides, for the first time, the seed transcriptome profile for the classified low grades (grade 2, and out-of-grade; i.e., grade 3) of rice under high-temperature stress condition.

Analyses of transgenic fibroblast growth factor 21 mature rice seeds.

Although some studies have been conducted on the effects of foreign protein expression on rice, the results vary with foreign gene types and protein expression. This study reveals the effects of fibroblast growth factor 21 (FGF21) expression on mature rice seeds in various aspects. Results revealed that the grain weight of the transgene rice was lower than that of non-transgenic wild-type. The sucrose content and ADP-glucose pyrophosphorylase (AGPase) activity in transgenic FGF21 rice were higher than that in non-transgenic wild-type rice, while changes in the starch content, starch branching enzyme (SBE), sucrose synthase (SuS), superoxide dismutase (SOD) and peroxidase (POD) activity were lower in transgenic FGF21 rice compared to non-transgenic wild-type. The scanning electron microscope results revealed that mature seeds of the transgenic FGF21 rice contained fewer vascular bundles with irregular arrangement compared to the wild-type. The mature seeds of CK and T1 rice lines were collected for proteome analysis, and 167 differentially expressed proteins (DEPs) were found. In addition, the most enriched pathways in both rice lines were determined to be amino sugar and nucleotide sugar metabolism and starch and sucrose metabolism, etc. This study laid the foundation for revealing the effects of exogenous protein expression on rice bioreactors.

Breeding of the Dormant Thermosensitive Genic Male-Sterile Lines of Early Rice to Overcome Pre-Harvest Sprouting of the Hybrid Seeds

The pollen development of male-sterile lines should be conducted during the high temperature season to ensure the sterility of male-sterile line pollen and the safety of hybrid seed production in the breeding of thermosensitive genic male-sterile (TGMS) lines of early rice. However, the mature hybrid rice seeds are prone to pre-harvest sprouting (PHS) during hot, rainy days, which leads to the decrease of hybrid rice seed quality or a loss of utilization value. In this study, with an aim to bring a dormancy trait from the dormant genotype to the current low-dormant TGMS lines, a common wild rice “Chaling” (Oryza rufipogon), which has strong seed dormancy, a large black stigma, and a high exsertion rate, was hybridized with the early rice TGMS line “Zhu 1S,” which often exhibits PHS. After screening for early maturing and seed dormancy, the early rice “Yezao S” and “Yezhuzao S,” with the characteristic of seed dormancy, were selected. The critical temperature of high temperature sterility for these two male-sterile lines was below 23.5 °C. The stigma of “Yezao S” was black and its exsertion rate was 79.9%, while “Yezhuzao S” had a colorless stigma with an exsertion rate of 76.8%. The expression of the genes OsNCEDs in “Yezao S” and “Yezhuzao S” were significantly higher than that in “Zhu 1S” the 15th day after heading, and the abscisic acid (ABA) content was also higher than that of “Zhu 1S” the 20th day after heading. The results indicated that introducing the dormancy characteristics from wild rice “Chaling” into the early rice TGMS lines was an effective way to prevent PHS.

Biolistic co-transformation of rice using gold nanoparticles

ABSTRACT- In order to produce transgenic rice lines lacking selectable marker gene, biolistic co-transformation technique using gold nanoparticles was adopted. In the first step, the efficiency of different sizes of gold particles was evaluated. The results showed that the efficiency of the nanoparticles in the transformation was comparable to that of the micro particles. Subsequently, two separate plasmids including p UBC harboring cry IA (c) gene and pTRA132 carrying hph gene as selectable marker gene were mixed and coated with gold nanoparticles. Embryo genic calli originated from mature rice seeds were targeted with plasmid-coated nan projectiles. Putative transgenic cells were selected from selection media after three subcultures in MS medium were supplemented with 50 mg hygromycine B l-1 in three weeks' intervals. The selected calli were then grown into putative transgenic plantlets. Molecular analysis performed on the regenerated lines showed that at least one copy of both genes was inserted into the genome of some transgenic lines. RT-PCR analysis also confirmed successful transcription of the cry IA (c) gene. Accordingly, it could be concluded that the biolistic co-transformation method using gold nanoparicles could be a successful method for gene delivery into rice varieties. Nevertheless, expression of cry IA (c) gene and loci of both genes should still be investigated in future studies.

Identification of ZHOUPI Orthologs in Rice Involved in Endosperm Development and Cuticle Formation.

The endosperm occupies most of the available space within mature rice seeds, contains abundant nutrients, and directly influences both the quality and quantity of rice production. Initial reports noted that AtZHOUPI (AtZOU) coordinates endosperm breakdown and the concomitant separation of the embryo from this structure in Arabidopsis. The results of this study show that rice genomes contain two most closely related homologs of AtZOU, OsZOU-1 and OsZOU-2; of these, OsZOU-1 expression is limited to within the endosperm where it can be detected throughout this structure 5 days after pollination (DAP). Its expression gradually decreases from seven DAP to nine DAP. The second of the two most closely related homologs, OsZOU-2, is highly expressed in leaves and stem, but is not detected in developing seeds. Heterologous expression of OsZOU-1 and OsZOU-2 in Atzou-4 mutants also revealed that OsZOU-1 partially complements the seed phenotypes of these individuals, while its counterpart, OsZOU-2, was unable to recover these phenotypes. The over-expression of OsZOU-1 severely disrupts both seed development and plant growth in transgenic rice lines, as plants in which this gene has been knocked down failed in the separation of endosperm from embryo and cuticle formation during seed development. The results of this study therefore suggest that OsZOU-1 is orthologous to the AtZOU, and regulates both endosperm development and cuticle formation in rice.

Application of whole sections of mature cereal seeds to visualize the morphology of endosperm cell and starch and the distribution of storage protein

The morphology of endosperm cell and starch and the distribution of storage protein affect the kernel weight and endosperm quality of cereal crops, and are of interest for crop breeders. They are significantly different in different regions of endosperm. Therefore, it is very important for in situ investigation of the morphology of endosperm cell and starch and the distribution of storage protein in mature cereal seed. In the present study, we reported a method for preparing the whole section of mature rice, maize, and wheat seeds. The section could be easily stained with fluorescent brightener 28, iodine solution, and coomassie brilliant blue R250 to exhibit the cell wall, starch, and storage protein. The observation of whole section of seed showed that the morphology of endosperm cell and starch and the distribution of storage protein were significantly different in different regions of endosperm of rice, maize, and wheat seed. The method could in situ investigate the morphology of endosperm cell and starch and the distribution of storage protein in not only mature cereal seed but also developing, germinated, and gelatinized cereal seed. This study was very useful for investigation of cereal seed development and utilization.

RNA-binding proteins associated with desiccation during seed development in rice

Long-lived mRNAs stored in mature seeds can remain active for long periods even if seeds undergo severe desiccation. They are then translated at the initiation of germination. To clarify the mechanism for stabilization of long-lived mRNAs during seed desiccation, fluctuations in RNA-binding protein (RBP) profiles that occur during seed formation in rice were analyzed. Proteomic analysis revealed that glycine-rich RBP 1A (GRP1A) is a highly abundant RBP in mature rice seeds. In addition, real-time RT-PCR analysis showed that putative RBP RZ-1A (RZ-1A) is seed specific. Moreover, transcripts of these two RBPs were clearly up-regulated during desiccation in rice seeds. The features of these two RBPs resemble those of late embryogenesis abundant proteins that function as molecular chaperones in dry seeds. Therefore, GRP1A and RZ-1A may have important roles in the stability of long-lived mRNAs in rice seeds.

Identification of Callus Induction Potential of Nine Different Genotypes of Indica Rice

Callus induction potential of nine economically important indica rice genotypes of Tamil Nadu and neighboring areas was examined on different callus induction medium. Nine genotypes include Improved White Ponni(I W Ponni), BPT-5204, TKM-9, CR1009, ADT-39, ADT-43, KKL(R) -1, CO(R)-48 and Savitri/ Ponmani. These rice genotypes were having distinct useful agronomical trait like resistance to various pest, tolerance to salinity and drought conditions. Most of the indica rice genotypes having good grain quality and nutritional properties are susceptible to pest and diseases causing loss in annual yield. The yield could be enhanced by genetic improvement by rice transformation to enable them to withstand biotic and abiotic stress. First step towards this approach is to produce a pleuripotent cell mass, callus induced from scutellum of mature rice seeds. MS basal medium with various 2, 4-Dichlorophenoxy acetic acid (2, 4-D) concentrations (2mg/L, 2.5mg/L, 3mg/L, 4mg/L and 5 mg/L) was used for these studies. Four indica rice genotypes I. W. Ponni, ADT- 43, BPT-5204 and TKM-9 found to be potent for callus induction from scutellum of mature seed at the concentration of 2mg/l to 5 mg /l 2,4-D. The varieties ADT-39, KKL (1) R, CO(R) 48, CR1009 and Savitri/Ponmani were recalcitrant and no embryogenic calli was obtained on either of the 2, 4-D concentration. 34 % of healthy calli obtained from I W Ponni 20 % of healthy callus induction obtained from BPT-5204 and 10 % of healthy calli from each of the ADT-43 and TKM-9. These indica rice genotypes could be used for various plant science researches.

High frequency plant regeneration from embryogenic callus of a popular indica rice (Oryza sativa L.).

The aim of the study is to establish a routine procedure for high frequency plant regeneration from in vitro raised embryogenic callus of abiotic salt sensitive indica rice (Oryza sativa L.) cultivar ADT 43. The effect of synthetic auxin 2,4-D on callus induction was optimized to achieve high frequency plant regeneration from fresh embryogenic callus without further subculture. Friable, nodular and creamish-white embryogenic callus cultures were raised from mature rice seeds on LS medium supplemented with 2.5 mg L(-1) 2,4-D and 1.0 mg L(-1) thiamine-HCL. Plant regeneration was achieved by the 24 days old embryogenic callus on MS medium supplemented with 1.0 mg L(-1) BAP and 1.5 mg L(-1)NAA. In vitro regenerated plants with multiple tillers and roots were transferred to sterile soil and maintained in the growth chamber. The regenerated plants exhibited normal growth and were phenotypically similar to plants maintained in the garden. Using the present protocol, 25-30 plantlets were regenerated from 50 individual mature seed derived callus within two to three months. This protocol has the potential for large-scale production of elite plants after genetic transformation.

Role of LOX3 Gene in Alleviating Adverse Effects of Drought and Pathogens in Rice

Lipoxygenase 3 (LOX3) is a major component of the LOX isozymes in mature rice seeds. To investigate the role of LOX3 gene under stresses, a plant expression vector containing antisense cDNA of LOX3 was constructed. Rice varieties Wuyunjing 7 and Kasalath were transformed by the Agrobacterium-mediated method and transgenic rice plants were generated. PCR and Southern blot results showed that the antisense LOX3 gene was integrated into the rice genome. Analyses of embryo LOX3 deletion and semi-quantitative RT-PCR confirmed the antisense suppression of LOX3 gene in transgenic plants. The T 2 antisense plants of LOX3 were sensitive to drought stress, rice blast and bacterial blight compared with non-transgenic plants. These results suggest that the LOX3 gene might function in response to stresses.

Posttranslational regulation of pyruvate, orthophosphate dikinase in developing rice (Oryza sativa) seeds

Pyruvate, orthophosphate dikinase (PPDK; E.C.2.7.9.1) is most well known as a photosynthetic enzyme in C4 plants. The enzyme is also ubiquitous in C3 plant tissues, although a precise non-photosynthetic C3 function(s) is yet to be validated, owing largely to its low abundance in most C3 organs. The single C3 organ type where PPDK is in high abundance, and, therefore, where its function is most amenable to elucidation, are the developing seeds of graminaceous cereals. In this report, we suggest a non-photosynthetic function for C3 PPDK by characterizing its abundance and posttranslational regulation in developing Oryza sativa (rice) seeds. Using primarily an immunoblot-based approach, we show that PPDK is a massively expressed protein during the early syncitial-endosperm/-cellularization stage of seed development. As seed development progresses from this early stage, the enzyme undergoes a rapid, posttranslational down-regulation in activity and amount via regulatory threonyl-phosphorylation (PPDK inactivation) and protein degradation. Immunoblot analysis of separated seed tissue fractions (pericarp, embryo + aleurone, seed embryo) revealed that regulatory phosphorylation of PPDK occurs in the non-green seed embryo and green outer pericarp layer, but not in the endosperm + aleurone layer. The modestly abundant pool of inactive PPDK (phosphorylated + dephosphorylated) that was found to persist in mature rice seeds was shown to remain largely unchanged (inactive) upon seed germination, suggesting that PPDK in rice seeds function in developmental rather than in post-developmental processes. These and related observations lead us to postulate a putative function for the enzyme that aligns its PEP to pyruvate-forming reaction with biosynthetic processes that are specific to early cereal seed development.

Effect of Genotype and Culture Media on Callus Induction and Plant Regeneration from Matured Rice Grain Culture

This study was under taken to elucidate the effect of genotypes and media compositions on callus induction from mature rice seeds. Three different callus induction media, designated as A (N6 mineral salts + N6 vitamins, 2 mg/l each + myoinositol, 100 mg/l + 2,4-D, 2.5 mg/l + kinetin, 0.5mg/l + AgNO3, 10 mg/l + maltose, 50 gm/l); B (MS organic salts + N6 mineral salts + NAA, 4 mg/l + kinetin, 2 mg/l + AgNO3, 5mg/l and sucrose, 60 gm/l); and C (B media without AgNO3), and six rice genotypes viz. Jumlimarshi, Tilki, Jethobudo, Manshara, Masuli and Pahenle were evaluated. The modified N6 medium supplemented with 2, 4-D, 2.5 mg/l and AgNO3, 10 mg/l exhibited better performance in callus induction. Among genotypes, callus induction frequency was higher (100%) in Masuli, Tilki and Jumlimarshi regardless of media tested. The positive effect of AgNO3 was only observed in medium A for quality callus induction and subsequent plant regeneration. The genotype Tilki performed better regarding plant regeneration (27.77%). Therefore, it is suggested that application of medium A is advantageous to accomplish overall efficiency of callus induction and plant regeneration from seeds of various rice genotypes. 
 
 Key words: 2, 4-D; AgNO3; dehulled rice; in vitro; regeneration 
 
 J. Inst. Agric. Anim. Sci. 26:21-26 (2005)

Enhanced Methionine and Cysteine Levels in Transgenic Rice Seeds by the Accumulation of Sesame 2S Albumin

A chimeric gene encoding a precursor polypeptide of sesame 2S albumin, a sulfur-rich seed storage protein, was expressed in transgenic rice plants under the control of the glutelin promoter with the aim of improving the nutritive value of rice. Rice grains harvested from the first generation of ten different transformed lines inherited the transgene, and the accumulated sesame 2S albumin was presumably processed correctly as its mature form in sesame seed. This transgene was specifically expressed in maturing rice seeds with its encoded sesame 2S albumin exclusively accumulated in the seeds. The crude protein content in rice grains from five putative homozygous lines was increased by 0.64-3.54%, and the methionine and cysteine contents of these transgenic rice grains were respectively elevated by 29-76% and 31-75% compared with those of wild-type rice grains.

The synergistic effects of sugar and abscisic acid on myo-inositol-1-phosphate synthase expression.

1L-myo-inositol-1-phosphate [Ins(1)P1] synthase (EC 5.5.1.4) catalyses the formation of Ins(1)P1 from glucose-6-phosphate, the first step in the biosynthesis of myo-inositol. Ins(1)P1 is a precursor of phytin (inositol hexakisphosphate), a storage form of phosphate and cations in seeds. Since sucrose and abscisic acid (ABA) are known to affect synthesis of storage compounds in seeds, we investigated the effects of ABA and sucrose on Ins(1)P1 synthase gene (RINO1) expression in cultured cells derived from the scutellum of mature rice seeds. Higher levels of RINO1 transcript accumulation were evident after treatment with either sucrose (10-100 mM) or ABA (10-8 M to 10-4 M). Glucose was also effective in the upregulation, whereas mannitol was not, suggesting that sucrose and glucose acted as metabolizable sugars and not as osmotica. Treatment with ABA and sucrose together resulted in much higher levels of transcript accumulation, suggesting a synergistic induction of the Ins(1)P1 synthase gene.