Identification of a New Giant Emrbryo Allele, and Integrated Transcriptomics and Metabolomics Analysis of Giant Embryo Development in Rice

Rice embryos are rich in high-quality protein, lipid, vitamins and minerals, representing the most important nutritional part of brown rice. However, the molecular mechanism of rice embryo development is poorly understood. In this study, two rice cultivars with contrasting embryo size (the giant embryo cultivar Dapeimi and the normal embryo cultivar 187R) were used to explore excellent genes controlling embryo size, and the developed near-isogenic lines (NILs) (NIL-D, which has the giant embryo phenotype, and its matching line, NIL-X) were used to explore transcript and metabolic properties in the earlier maturation stage of giant embryo development under natural conditions. The map-based cloning results demonstrated that Dapeimi is a novel allelic mutant of the rice GIANT EMBRYO (GE) gene, and the functional mutation site is a single cytosine deletion in the exon1. A total of 285 differentially accumulated metabolites (DAMs) and 677 differentially expressed genes (DEGs) were identified between NIL-D and NIL-X. The analysis of DAMs indicated that plants lacking GE mainly promoted energy metabolism, amino acid metabolism, and lipid metabolism pathways in the rice embryo. Pearson correlation coefficient showed that 300 pairs of gene-metabolites were highly correlated. Among them, OsZS_02G0528500 and OsZS_12G0013700 were considered to be key genes regulating L-Aspartic acid and L-Tryptophan content during rice giant embryo development, which are promising to be good candidate genes to improve rice nutrition. By analyzing rice embryo development through a combination of strategies, this research contributes to a greater understanding of the molecular mechanism of rice embryo development, and provides a theoretical foundation for breeding high-nutrition varieties.

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Improvement of soybean somatic embryo development and maturation by abscisic acid treatment

Canadian Journal of Plant Science ◽

10.4141/p99-056 ◽

2000 ◽

Vol 80 (2) ◽

pp. 271-276 ◽

Cited By ~ 10

Author(s):

Lining Tian ◽

Daniel C. W. Brown

Keyword(s):

Somatic Embryogenesis ◽

Abscisic Acid ◽

Embryo Development ◽

Development Stage ◽

Normal Embryo ◽

Maturation Stage ◽

Globular Stage ◽

Solid Media ◽

Stage Embryo

Recovery of tissue culture-derived plants through somatic embryogenesis is a useful system for genetic engineering of soybean. The effect of abscisic acid (ABA) on soybean somatic embryogenesis, development, and maturation was investigated. ABA at 1, 10, 50, 100, and 500 µM were applied at different stages of embryo development; namely, at the globular stage in suspension culture, at the development stage and at the maturation stage on solid media. ABA promoted embryo growth and development when applied at the globular stage. Embryo size, after 15 d and after 1 mo on development medium, was significantly greater than that without exposure to ABA. ABA promoted normal embryo morphogenesis and 62% more normal embryos developed when embryos were treated with ABA at the globular stage. ABA treated-embryos showed an increased tolerance to partial desiccation (from 24% to 78%) and exhibited an increased germination capability relative to non-ABA-treated controls (54% versus 8%). Somatic embryos appeared to undergo a decreasing sensitivity to ABA during maturation. ABA did not show an effect when applied during embryo development and maturation stages. A protocol for more normal embryo formation and improved embryo germination is reported. Key words: Glycine max, somatic embryogenesis, in vitro culture

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Somatic Cell Nuclear Transfer Embryos Show Massive Dysregulation of Genes Involved in Transcription Pathway

10.1101/2020.12.06.413393 ◽

2020 ◽

Author(s):

Chunshen Long ◽

Hanshuang Li ◽

Xinru Li ◽

Yongchun Zuo

Keyword(s):

Embryo Development ◽

Nuclear Transfer ◽

Normal Embryo ◽

Cloning Efficiency ◽

Rna Seq ◽

Activation Of Transcription ◽

Potential Association ◽

Gene Regulatory ◽

Genome Activation ◽

Zygotic Genome

AbstractTranscription is the most fundamental molecular event that occurs with zygotic genome activation (ZGA) during embryo development. However, the potential association between transcription pathways and low cloning efficiency of nuclear transfer (NT) embryos remains elusive. Here, we integrated a series of RNA-seq data on NT embryos to deciphering the molecular barriers of NT embryo development. Comparative transcriptome analysis indicated that incomplete activation of transcription pathways functions as a barrier for NT embryos. Then, the gene regulatory network (GRN) identified that crucial factors responsible for transcription play a coordinated role in epigenome erasure and pluripotency regulation during normal embryo development. But in NT embryos, massive genes involved in transcription pathways were varying degrees of inhibition. Our study therefore provides new insights into understanding the barriers to NT embryo reprogramming.

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Control of Rice Embryo Development, Shoot Apical Meristem Maintenance, and Grain Yield by a Novel Cytochrome P450

Molecular Plant ◽

10.1093/mp/sst107 ◽

2013 ◽

Vol 6 (6) ◽

pp. 1945-1960 ◽

Cited By ~ 37

Author(s):

Weibing Yang ◽

Mingjun Gao ◽

Xin Yin ◽

Jiyun Liu ◽

Yonghan Xu ◽

...

Keyword(s):

Cytochrome P450 ◽

Grain Yield ◽

Embryo Development ◽

Shoot Apical Meristem ◽

Apical Meristem ◽

Meristem Maintenance ◽

Rice Embryo

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Styrene impairs normal embryo development in the Mediterranean mussel (Mytilus galloprovincialis)

Aquatic Toxicology ◽

10.1016/j.aquatox.2018.05.026 ◽

2018 ◽

Vol 201 ◽

pp. 58-65 ◽

Cited By ~ 7

Author(s):

Rajapaksha Haddokara Gedara Rasika Wathsala ◽

Silvia Franzellitti ◽

Morena Scaglione ◽

Elena Fabbri

Keyword(s):

Embryo Development ◽

Mytilus Galloprovincialis ◽

Normal Embryo ◽

Mediterranean Mussel ◽

The Mediterranean

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Kinetics of Douglas-fir (Pseudotsuga menziesii) somatic embryo development

Canadian Journal of Botany ◽

10.1139/b98-050 ◽

1998 ◽

Vol 76 (5) ◽

pp. 863-871 ◽

Cited By ~ 2

Author(s):

Ryan P Taber ◽

Chun Zhang ◽

Wei-Shou Hu

Keyword(s):

Somatic Embryo ◽

Embryo Development ◽

Pseudotsuga Menziesii ◽

Growth Kinetics ◽

Selection Process ◽

Scale Up ◽

Douglas Fir ◽

Maturation Stage ◽

Mature Embryos ◽

Maintenance Stage

Douglas-fir (Pseudotsuga menziesii (Mirb.) Franco) is one of the most economically important softwood species in the Pacific Northwest region. Somatic embryogenesis is a potential mass propagation technology for increasing the productivity of existing forest acreage. Combined with traditional breeding methods and recent advances of genetic engineering in plant species, somatic embryos can shorten the elite clone selection process significantly. Somatic embryo culture of Douglas-fir involves three stages: maintenance, abscisic acid (ABA) singulation, and maturation. At the beginning of all stages of culture, the population of cells with embryogenic potential is increased through the weekly subcultured maintenance stage; transfer into the ABA singulation stage initiates embryo development, while cotyledonary embryos are formed in the maturation state. The first two stages were carried out in submerged suspension culture, while during the maturation stage the developing embryos were placed on a polyester pad in a Petri dish. The growth kinetics in these stages were investigated. Fresh and dry weights were observed to double in the maintenance stage, while a smaller increase occurred in the ABA singulation stage. NH4+ was consumed preferentially to NO3- in all culture stages. Sucrose, the primary carbon source, was hydrolyzed to glucose and fructose rapidly. During cultivation, glucose and fructose were consumed simultaneously. The hydrolysis of sucrose resulted in a slight osmolarity increase at the beginning of all culture stages, while the subsequent consumption of glucose and fructose coincided with a gradual decrease in osmolarity. This dynamic osmolarity pressure profile is most profound in the maturation stage, in which the initial high osmotic pressure of 600 ± 20 mOsm/kg (mean ± SD) increased to 700 ± 27 mOsm/kg after sucrose was hydrolyzed but decreased to 350 ± 14 mOsm/kg after the depletion of sugars at the end of cultivation. The complete process of embryo development, from the week-long maintenance culture, through the weekly subcultured ABA singulation culture, to the maturation of embryos took between 70 and 80 days. Each millilitre of culture present at the onset of maintenance culture gave rise to approximately 420 mature embryos. During that same time period, the biomass increased approximately 100 times. Prolonging the cultivation time failed to increase the yield of mature embryos. These results give a more complete view of the kinetic behavior of developing Douglas-fir embryos and will aid in the optimization and scale-up of this important process.Key words: somatic embyrogenesis, Douglas-fir, Pseudotsuga menziesii, growth kinetics, ABA, osmolarity, development.

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Mitochondria and vesicles differ between adult and prepubertal sheep oocytes during IVM

Reproduction Fertility and Development ◽

10.1071/rd13359 ◽

2015 ◽

Vol 27 (3) ◽

pp. 513 ◽

Cited By ~ 12

Author(s):

Karen L. Reader ◽

Neil R. Cox ◽

Jo-Ann L. Stanton ◽

Jennifer L. Juengel

Keyword(s):

Embryo Development ◽

Quantitative Polymerase Chain Reaction ◽

Developmental Competence ◽

Normal Embryo ◽

Chain Reaction ◽

Mitochondrial Dna Copy Number ◽

Storage Vesicle ◽

Mitochondrial Volume ◽

Polymerase Chain ◽

Mitochondrial Number

Oocytes from prepubertal animals have a reduced ability to undergo normal embryo development and produce viable offspring. The correct quantity, activity and cytoplasmic distribution of oocyte organelles are essential for oocyte maturation, fertilisation and subsequent embryo development. The aim of this study was to quantify the ultrastructural differences between oocytes from prepubertal lamb and adult ewes using electron microscopy and stereology. We also determined whether quantitative polymerase chain reaction (qPCR) methods give comparable estimates of mitochondrial number to stereology. Mean storage vesicle volume was greater in adult compared with lamb oocytes before IVM and decreased during maturation in both adult and lamb oocytes. Mitochondrial volume and number increased in adult oocytes during maturation; however, no increase was observed in lamb oocytes. Mitochondrial DNA copy number measured by qPCR showed no differences between adult and lamb oocytes. A different distribution of mitochondria was observed in lamb oocytes before maturation, while the percentage of hooded mitochondria increased during maturation in adult oocytes and decreased in the lamb. In conclusion, the present study has identified differences in the vesicles and mitochondria between adult and lamb oocytes from ewes that may contribute to reduced developmental competence in prepubertal oocytes.

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99 Short-term storage of equine embryos at 5 or 20°C does not cause lipid peroxidation

Reproduction Fertility and Development ◽

10.1071/rdv32n2ab99 ◽

2020 ◽

Vol 32 (2) ◽

pp. 175

Author(s):

G. D. Gastal ◽

D. Scarlet ◽

C. Aurich

Keyword(s):

Lipid Peroxidation ◽

Embryo Development ◽

Storage Temperature ◽

Storage Period ◽

Small Sample ◽

Temperature Curve ◽

Data Logger ◽

Limit Of Quantification ◽

Embryo Size ◽

Holding Temperature

Maintaining the integrity of equine embryos during storage for transportation is essential for successful conception after transfer. During storage, reactive oxygen species may originate from embryo metabolism, causing lipid peroxidation and increasing its end product malondialdehyde (MDA); MDA is one of the important biomarkers for oxidative stress. This study aimed to evaluate the temperature curve, pH and lipid peroxidation of equine embryos stored in holding medium after 24h at 5 or 20°C. Embryos (n=33) were collected on Day 7 (n=21, 7 embryos per group) or Day 8 (n=12) after ovulation and assigned to four groups: Day 7 control (D7, fresh); Day 7, 24h at 5°C (E5C); Day 7, 24h at 20°C (E20C); and Day 8 control (D8, fresh 24h time control). After collection, embryos were washed and kept in holding medium (Minitube) for morphological classification and measurements. For pH and lipid peroxidation measurement, embryos were kept in a fixed volume of holding medium (150µL) within a microtube (200µL); the microtube was kept within a falcon tube (50mL) inside of an Equitainer (Hamilton Biovet). The temperature was recorded by a data logger (Testo 175, Testo) every 10min for 24h. The pH was assessed by a pH meter using a microelectrode (InLab Ultra-Micro-ISM, Mettler Toledo) for small sample volumes. Lipid peroxidation was assessed using the MDA assay kit (catalog number MAK085, Sigma-Aldrich) according to the manufacturer's instructions. Statistical analyses were performed using the Kruskal-Wallis nonparametric test and Mann-Whitney U to compare differences among groups. Embryo size differed (P<0.05) between D7 (383±41 µm) and D8 (1044±131 µm). Storage temperature (E5C or E20C) did not affect embryo size (382±47 and 553±99µm, respectively; P>0.05). The temperature curve was similar (P>0.05) among embryos within the treatment groups during the storage period. The pH (7.22±0.07 and 7.22±0.09) did not differ (P>0.05) between E5C and E20C. Lipid peroxidation levels were below the limit of quantification (0.04 nmol) in all groups. The present findings suggest that holding temperature does not affect the size, pH, or lipid peroxidation of equine embryos stored for 24h in holding medium. However, our previous studies (Gastal et al. 2018 J. Equine Vet. Sci. 66, 185) have shown that holding temperature influences the expression of genes involved in equine embryo development. In conclusion, variations regarding embryo development and conception rate after transfer of embryos stored at different temperatures might be related to factors other than lipid peroxidation.

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257. Activation of a calcium-activated chloride channel by paf is required for normal preimplantation mouse embryo development in vitro

Reproduction Fertility and Development ◽

10.1071/srb08abs257 ◽

2008 ◽

Vol 20 (9) ◽

pp. 57

Author(s):

Y. Li ◽

M. L. Day ◽

C. O.'Neill

Keyword(s):

Embryo Development ◽

Mouse Embryo ◽

Preimplantation Embryo ◽

Platelet Activating Factor ◽

Outward Current ◽

Normal Embryo ◽

Cell Stage ◽

Preimplantation Embryo Development ◽

Cl Channel

Platelet activating factor (paf) is an autocrine survival factor for preimplantation embryo. Binding of paf to its receptor activates PI3kinase, causing an IP3-dependent release of Ca2+ from intracellular stores as well as activation of Ca2+ influx via a dihydropyridine-sensitive Ca2+ channel. These actions result in the generation of a defined intracellular calcium ([Ca2+]i) transient in the 2-cell embryo[1]. By using combined whole-cell patch-clamp and real-time [Ca2+]i analyses, we have shown that paf also induces a concomitant hyperpolarisation of the membrane potential in 2-cell embryos, accompanied by an increased net outward ion current. Both the membrane hyperpolarisation and outward current were dependent upon the occurrence of the paf-induced [Ca2+]i transient[2]. The aim of this study was to investigate the characteristics of the paf-induced outward current in 2-cell embryos and to assess whether it has a role in normal mouse preimplantation development. We show that: (1) removal of extracellular anions or treatment with niflumic acid (NFA, 100 μM, a Ca2+-activated Cl- channel blocker) prevented activation of the outward current by paf but had no effect on the paf-induced [Ca2+]i transient; and (2) The culture of embryos with NFA (100 μM) from the 1-cell to late 2-cell stage significantly reduced their development to the blastocyst stage (P < 0.001), but treatment with NFA from the late 2-cell stage had no effect on development. The results show that paf induces an increase in [Ca2+]i which in turn activates a Ca2+-activated Cl- channel. The activity of this NFA-sensitive channel during the zygote to 2-cell stage is required for normal embryo development. (1) C. O’Neill (2008) The potential roles of embryotrophic ligands in preimplantation embryo development. Hum Reprod Update 14:275–288. (2) Y. Li, M.L. Day & C. O’Neill (2007) Autocrine activation of ions currents in the two-cell mouse embryo. Exp Cell Res. 313:2785–2794.

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