Embryonic Axis Research Articles

Tailored Media Are Key to Unlocking the Diversity of Endophytic Bacteria in Distinct Compartments of Germinating Seeds.

ABSTRACTSeeds offer an internal microbial niche, termed the endosphere, colonized by communities of endophytic bacteria. To elucidate the functions of seed endophytes during germination and early plant growth, studies with culturable isolates are essential. Conventional growth media favor few fast-growing taxa, while micro organisms with restricted nutrient requirements are usually outcompeted prior to isolation. Consequently, current knowledge of the interaction between seeds and their endophytes remains limited to only few bacterial taxa, despite a “black box” of unculturable isolates colonizing the endosphere. Here, we designed various solid media to mimic the endosphere of germinating soybean (Glycine max L.) seeds and assessed their effect on the diversity of culturable endophytic bacteria. The embryonic axis (i.e., the future plant) possessed higher richness and harbored more unique genera (i.e., Brevundimonas, Methylobacterium, Microbacterium, Pseudoclavibacter, and Rathayibacter) than cotyledons (i.e., seed storage organs). Overall, media containing germinating and ground seeds enabled culturing and isolation of the broadest diversity of endophytic bacteria, viewed through the molecular identification of 246 isolates. The use of multiple tailored media helped uncover trophic adaptation of the core taxa. Furthermore, comparison of seeds from four lots of distinct cultivars and origin revealed few overlapping taxa, indicating that the parental environment, including soil and fertilization regime, influenced seed endophytic diversity. Extended diversity of native seed endophytic bacteria revealed the functional relevance of unique Arthrobacter, Bacillus, and Curtobacterium strains to seed germination under salt stress, exemplifying the importance of enhanced culturing approaches to elucidate the role of microbiota in seed germination.IMPORTANCE Plant growth-promoting endophytic isolates that appear to advance seed germination are often obtained from plant niches other than the seed endosphere. Isolating pure cultures of native endophytes from seeds during germination is crucial to investigate their function during early plant growth. Here, the diversity of endophytic bacteria isolated from seeds during soybean germination was enhanced by combining media tailored to the nutritional composition of the seed endosphere, including pregerminated seeds themselves. Our results show that isolation from distinct soybean seed compartments affected such diversity, with the embryonic axis harboring more unique taxa while displaying higher endophytic richness. Furthermore, using pools of seeds from separate lots, each corresponding to a certain cultivar and field site, supported isolation of further unique strains that often unveiled substantial effects on germination performance. Such findings are relevant to assist studies on the interactions between seeds and their native endophytic bacteria.

Clue of zinc oxide and copper oxide nanoparticles in the remediation of cadmium toxicity in Phaseolus vulgaris L. via the modulation of antioxidant and redox systems.

The present study represents new evidence of the successful use of metal oxide nanoparticles in the remediation of heavy metals. Zinc oxide nanoparticles (ZnO NP) and copper oxide nanoparticles (CuO NP) were assessed to alleviate cadmium (Cd) toxicity in Phaseolus vulgaris L. seedlings and plants. Monitoring physiological and metabolic parameters allowed to elucidate Cd mechanism and process whereby it exerts phytotoxic effects on bean. The response of P. vulgaris seedlings is NP dose-dependent (10 mg/L, 50 mg/L, 100 mg/L, and 200 mg/L). Similarly, applied concentrations triggered a differential response of growing plants in terms of length and biomass. Our physiological data allowed to select 100 mg/L as the most appropriate concentration to apply, in order to avoid any risk of phytotoxicity. The regulatory mechanisms by which ZnO NP and CuO NP act are for the first time compared in the embryonic axes of bean seedlings under Cd stress. Both NP were able to reduce the hypergeneration of hydrogen peroxide (H2O2). They also acted via enhancing ROS scavenging enzymatic capacity, and activity of antioxidant enzymes CAT, APX, GPOX, GPX, and GR, and inhibited the activity of ROS producing enzymes such as GOX and NOX. Another mechanistic effect of NP consisted of the modulation of redox enzymes Trx, NTR, Fd, and FNR evolved in cellular homeostasis and maintaining reduced status in cells. Taken together, ZnO NP triggered more significant metabolic regulations allowing to mitigate the oxidative damage caused by Cd.

Seed-to-Seedling Transition in Pisum sativum L.: A Transcriptomic Approach.

The seed-to-seedling transition is a crucial step in the plant life cycle. The transition occurs at the end of seed germination and corresponds to the initiation of embryonic root growth. To improve our understanding of how a seed transforms into a seedling, we germinated the Pisum sativum L. seeds for 72 h and divided them into samples before and after radicle protrusion. Before radicle protrusion, seeds survived after drying and formed normally developed seedlings upon rehydration. Radicle protrusion increased the moisture content level in seed axes, and the accumulation of ROS first generated in the embryonic root and plumule. The water and oxidative status shift correlated with the desiccation tolerance loss. Then, we compared RNA sequencing-based transcriptomics in the embryonic axes isolated from pea seeds before and after radicle protrusion. We identified 24,184 differentially expressed genes during the transition to the post-germination stage. Among them, 2101 genes showed more prominent expression. They were related to primary and secondary metabolism, photosynthesis, biosynthesis of cell wall components, redox status, and responses to biotic stress. On the other hand, 415 genes showed significantly decreased expression, including the groups related to water deprivation (eight genes) and response to the ABA stimulus (fifteen genes). We assume that the water deprivation group, especially three genes also belonging to ABA stimulus (LTI65, LTP4, and HVA22E), may be crucial for the desiccation tolerance loss during a metabolic switch from seed to seedling. The latter is also accompanied by the suppression of ABA-related transcription factors ABI3, ABI4, and ABI5. Among them, HVA22E, ABI4, and ABI5 were highly conservative in functional domains and showed homologous sequences in different drought-tolerant species. These findings elaborate on the critical biochemical pathways and genes regulating seed-to-seedling transition.

Evaluation of transgenic chickpea harboring codon-modified Vip3Aa against gram pod borer (Helicoverpa armigera H.).

The gram pod borer is a major pest of chickpea, accounting for average annual yield losses to the tune of 40–50%. VIP3Aa, a class of insecticidal protein with different receptor binding site in the insect’s midgut compared to Bt-crystal protein, offers an alternative protection strategy against Lepidopteran insects. Here, we report evaluation of genetically engineered chickpea lines harboring codon modified Vip3Aa (cmVip3Aa) against the Lepidopteran insect pest, gram pod borer. The synthetic codon modified, cmVip3Aa gene of 2,370 bp was sub-cloned in modified plant expression vector and used for direct transformation of embryonic axis explants of chickpea (cv. DCP 92–3), with transformation efficiency of 4.30%. Presence and transmission of transgene across two generations were confirmed by PCR and Southern blot analyses in the five selected transgenic chickpea lines. Real Time PCR analyses indicated variable levels of cmVip3Aa expression in the transgenic chickpea lines (average Cq values 15.01±0.86 to 19.32±0.10), which were absent in the non-transgenic counterpart. Detached leaf insect bioassay indicate larval mortality (up to 39.75%), reduced larval feeding (up to 82.91%) and reduced larval weight gain (up to 68.23%), compared to control lines. Evaluation of gene offers a platform to identify efficacious insecticidal gene that can be used for insect resistance management in chickpea.

Genetic transformation of common beans (Phaseolus vulgaris L.) through Agrobacterium tumefaciens carrying Cry1Ab gene.

Seed beetles are one of the most important causes of yield loss in bean production. It is essential to develop resistant varieties in the fight against these pests. Agrobacterium-based gene transformation is the most widely used breeding method worldwide to develop insect-resistant varieties. Embryonic axes and plumule explants were obtained from Agrobacterium tumefciens treated mature zygotic embryos of low and high raw protein-based common bean cultivars Akman 98 and Karacaşehir 90. Agrobacterium tumefaciens contained a synthetic Bacillus thuringiensis insecticidal crystal protein gene (Bt Cry1Ab) controlled by the 35S promoter and NOS terminator sequences. The transformation event was genotype and explant dependent. The plumule explants could not withstand kanamycin-based selection pressure and died. It was possible to get two transgenic plants using embryonic axis explants of low protein cultivar Akman 98. These results were validated using GUS analysis, PCR, RT-PCR, bioassay analysis, and ELISA test from the samples taken from T0 and T1 generations. Bioassay tests showed that these plants were protected from the damage of legume seed insects (Bruchus spp.). The results are very encouraging and may help in producing better transgenic common bean germplasm leading to safe agriculture and reducing environmental pollutions.

Gene Expression Analysis of the Stress Response to Lithium, Nickel, and Zinc in Paracentrotus lividus Embryos.

Many anthropogenic pollutants such as metals are discharged into the marine environment through modern sources. Among these, lithium (Li), nickel (Ni), and zinc (Zn) can interfere with biological processes in many organisms when their concentration rises. These metals are toxic to sea urchin embryos, affecting their development. Indeed, animal/vegetal and dorso/ventral embryonic axes are differently perturbed: Li is a vegetalizing agent, Ni can disrupt dorso-ventral axis, Zn can be animalizing. To address the molecular response adopted by embryos to cope with these metals or involved in the gene networks regulating embryogenesis, and to detect new biomarkers for evaluating hazards in polluted environments in a well-known in vivo model, we applied a high-throughput screening approach to sea urchin embryos. After fertilization, Paracentrotus lividus embryos were exposed to Li, Ni, and Zn for 24/48 h. At both endpoints, RNAs were analyzed by NanoString nCounter technology. By in silico analyses, we selected a panel of 127 transcripts encoding for regulatory and structural proteins, ranked in categories: Apoptosis, Defense, Immune, Nervous, Development, and Biomineralization. The data analysis highlighted the dysregulation of many genes in a metal-dependent manner. A functional annotation analysis was performed by the KEEG Orthology database. This study provides a platform for research on metals biomarkers in sea urchins.

Multi-Omics Approaches Unravel Specific Features of Embryo and Endosperm in Rice Seed Germination.

Seed germination and subsequent seedling growth affect the final yield and quality of the crop. Seed germination is defined as a series of processes that begins with water uptake by a quiescent dry seed and ends with the elongation of embryonic axis. Rice is an important cereal crop species, and during seed germination, two tissues function in a different manner; the embryo grows into a seedling as the next generation and the endosperm is responsible for nutritional supply. Toward understanding the integrated roles of each tissue at the transcriptional, translational, and metabolic production levels during germination, an exhaustive “multi-omics” analysis was performed by combining transcriptomics, label-free shotgun proteomics, and metabolomics on rice germinating embryo and endosperm, independently. Time-course analyses of the transcriptome and metabolome in germinating seeds revealed a major turning point in the early phase of germination in both embryo and endosperm, suggesting that dramatic changes begin immediately after water imbibition in the rice germination program at least at the mRNA and metabolite levels. In endosperm, protein profiles mostly showed abundant decreases corresponding to 90% of the differentially accumulated proteins. An ontological classification revealed the shift from the maturation to the germination process where over-represented classes belonged to embryonic development and cellular amino acid biosynthetic processes. In the embryo, 19% of the detected proteins are differentially accumulated during germination. Stress response, carbohydrate, fatty acid metabolism, and transport are the main functional classes representing embryo proteome change. Moreover, proteins specific to the germinated state were detected by both transcriptomic and proteomic approaches and a major change in the network operating during rice germination was uncovered. In particular, concomitant changes of hormonal metabolism-related proteins (GID1L2 and CNX1) implicated in GAs and ABA metabolism, signaling proteins, and protein turnover events emphasized the importance of such biological networks in rice seeds. Using metabolomics, we highlighted the importance of an energetic supply in rice seeds during germination. In both embryo and endosperm, starch degradation, glycolysis, and subsequent pathways related to these cascades, such as the aspartate-family pathway, are activated during germination. A relevant number of accumulated proteins and metabolites, especially in embryos, testifies the pivotal role of energetic supply in the preparation of plant growth. This article summarizes the key genetic pathways in embryo and endosperm during rice seed germination at the transcriptional, translational, and metabolite levels and thereby, emphasizes the value of combined multi-omics approaches to uncover the specific feature of tissues during germination.

Preservation in a cryobank of Juglans regia L. accession from several populations in the Sairam-Ugam State National Natural Park

Persian walnut (Juglans regia L.) is one of the most economically important nut crops. In Kazakhstan, the wild-growing population of Juglans regia is registered in the south of the country and is protected on the territory of the Sairam-Ugam State National Natural Park. As a result of expeditions in 2018-2019 136 accessions of Juglans regia were described according to the descriptors of the International Plant Genetic Resources Institute, fruits of 73 accessions of walnuts were collected, which are highly diverse in terms of size, shape and weight of nuts, kernel yield, color and texture of the shell and other parameters. To conserve the biodiversity of Juglans regia, the efficient cryopreservation technique has been developed for isolated embryonic axes. After surface disinfection with 0.1% mercuric chloride solution and rinsing with sterile distilled water, the embryonic axes were desiccated under laminar flow 1 hour to moister content of 12.1%, placed in cryovials, and immersed in a Dewar vessel with liquid nitrogen. In 72.4% of embryonic axes after cryogenic storage, plant recovery was noted on the Driver-Kuniyuki walnut medium with 1 mg/l 6-benzylaminopurine and 0.01 mg/l indolylbutyric acid. A cryogenic collection of 73 Juglans regia accessions has been established. For the first time, the genetic material of Juglans regia, growing in Kazakhstan, has been stored in a cryogenic bank at -196°C. Key words: Juglans regia L., walnut, cryopreservation, cryobank, isolated embryonic axes

Viability and vigor of Moringa oleifera Lam. seeds using the tetrazolium test

To evaluate the physiological potential of the seeds, the tetrazolium test would be an interesting alternative, as it is a quick test that provides a detailed analysis of viability and vigor. Therefore, it is necessary to define pre-conditioning conditions, such as temperature and duration of soaking, concentration of the tetrazolium solution, and method of exposing the seeds to staining. Moringa oleifera Lam., belonging to the family Moringaceae, is widely distributed in tropical countries and easily adapted to semi-arid conditions, possessing medicinal value, as forage, condiment, in the cosmetics industry, melliferous, fuel, and in water purification treatment through the coagulant solution made from seeds. Based on the above, the objectives of the present work were to establish the most appropriate procedure for conducting the tetrazolium test to evaluate the viability and vigor of M. oleifera seeds. Only the cotyledons containing the embryo were placed in 50 mL plastic cups and immersed in a solution of 2,3,5-triphenyl chloride tetrazolium salt at concentrations of 0.05; 0.075 and 0.1% for staining periods of 1, 2, and 3 hours in a Biochemical Oxygen Demand chamber at 30 ºC in the dark. The seeds were evaluated one by one for tissue condition (firm or flaccid) and seed color, and by the location and size of the lesions on the cotyledon and embryonic axis. For the standardization of the tetrazolium test, the Completely Randomized Design was used in a 3x3+1 factorial scheme (three concentrations of tetrazolium solution x three staining periods + control = germination test) with four repetitions of 25 seeds. M. oleifera seeds were found to have a high germination rate (100%), there was no significant difference between the control and the percentage of viable seeds (TZ) for the 2 and 3 hours staining periods, immersed in tetrazolium solution at concentrations of 0.075 (96%) and 0.05% (99%), respectively. The most appropriate procedure for conducting the tetrazolium test to assess the viability and vigor of M. oleifera seeds is to use a concentration of 0.05% for 3 hours of salt immersion

Guidance by followers ensures long-range coordination of cell migration through α-catenin mechanoperception.

Morphogenesis, wound healing, and some cancer metastases depend upon the migration of cell collectives that need to be guided to their destination as well as coordinated with other cell movements. During zebrafish gastrulation, the extension of the embryonic axis is led by the mesendodermal polster that migrates toward the animal pole, followed by the axial mesoderm that undergoes convergence and extension. Here, we investigate how polster cells are guided toward the animal pole. Using a combination of precise laser ablations, advanced transplants, and functional as well as in silico approaches, we establish that each polster cell is oriented by its immediate follower cells. Each cell perceives the migration of followers, through E-cadherin/α-catenin mechanotransduction, and aligns with them. Therefore, directional information propagates from cell to cell over the whole tissue. Such guidance of migrating cells by followers ensures long-range coordination of movements and developmental robustness.

Cold stratification-induced dormancy removal in apple (Malus domestica Borkh.) seeds is accompanied by an increased glutathione pool in embryonic axes

A reduced form of glutathione (GSH) is an essential metabolite that participates in the control of reactive oxygen species (ROS) levels in cells. GSH plays a pivotal role in seed biology as a modulator of seed viability and germination. The GSH:GSSG ratio and half-cell reduction potential (EGSSG/2GSH) serve as indicators of the oxidative status in seeds. Apple (Malus domestica Borkh.) seeds are deeply dormant, and this state is removed by long-term cold stratification. The aim of our work was to examine the modification of GSH and GSSG content, GSH:GSSG ratio and EGSSG/2GSH in the embryonic axes isolated from apple seeds subjected to cold stratification for 7, 14, 21 and 40 d. Our data indicated that cold stratification increased the generation of free radicals in the embryonic axes, which correlated with an alteration in the expression of genes encoding Rboh, particularly RbohC. GSH and GSSG levels increased during prolonged cold stratification of apple seeds. This was accompanied by the modification of glutathione reductase and glutathione peroxidase-like activities, which did not match their transcript levels. The steady-state GSH:GSSG ratio and EGSSG/2GSH in the axes of embryos subjected to cold stratification indicated no impact of the dormancy removal treatment on apple seed viability. We suggest that the glutathione system is an important component of the redox network and is involved in the management of the seed transition from dormant to nondormant states.

Epigenetic Marks, DNA Damage Markers, or Both? The Impact of Desiccation and Accelerated Aging on Nucleobase Modifications in Plant Genomic DNA.

Modifications of DNA nucleobases are present in all forms of life. The purpose of these modifications in eukaryotic cells, however, is not always clear. Although the role of 5-methylcytosine (m5C) in epigenetic regulation and the maintenance of stability in plant genomes is becoming better understood, knowledge pertaining to the origin and function of oxidized nucleobases is still scarce. The formation of 5-hydroxymetylcytosine (hm5C) in plant genomes is especially debatable. DNA modifications, functioning as regulatory factors or serving as DNA injury markers, may have an effect on DNA structure and the interaction of genomic DNA with proteins. Thus, these modifications can influence plant development and adaptation to environmental stress. Here, for the first time, the changes in DNA global levels of m5C, hm5C, and 8-oxo-7,8-dihydroguanine (8-oxoG) measured by ELISA have been documented in recalcitrant embryonic axes subjected to desiccation and accelerated aging. We demonstrated that tissue desiccation induces a similar trend in changes in the global level of hm5C and 8-oxoG, which may suggest that they both originate from the activity of reactive oxygen species (ROS). Our study supports the premise that m5C can serve as a marker of plant tissue viability whereas oxidized nucleobases, although indicating a cellular redox state, cannot.

Ectopic expression of Vigna radiata's vacuolar Na+/H+ antiporter gene (VrNHX1) in indica rice (Oryza sativa L.)

It is essential to develop high salt-tolerant rice varieties in order to cultivate the salt-affected lands. In this study, Na+/H+ exchanger 1 (NHX1) gene isolated from Vigna radiata L. Wilczek was transferred in Bangladesh Rice Research Institute (BRRI) developed two indica rice genotypes BRRI Dhan28 and BRRI Dhan29 using in-planta approach for improvement of salinity tolerance. Embryonic axes of matured dehusked rice seeds were injured and co-cultivated with Agrobacterium strain harboring VrNHX1 gene and finally regenerated. GUS histochemical assay and PCR amplification of GUS-a and VrNHX1 were performed to confirm the transformation. Expression confirmation was done by semi-quantitative RT-PCR. Under salinity stress, transgenic lines showed higher chlorophyll, relative water content and decreased electrolyte leakage, proline content, lipid peroxidation level, and catalase enzyme activity which represent the better physiology than control plants. Moreover, under salinity stress (150 mM), transgenic lines exhibited superior growth and salt tolerant than non-transgenic plants.

The Enigmatic Aliphatic Acetogenins and Their Correlations With Lipids During Seed Germination and Leaf Development of Avocado (Persea americana Mill.).

Lipids in avocados have been widely studied due to their nutritional value and several reported bioactivities. Aliphatic acetogenins are a relevant component of the avocado lipidome and have been tested for several potential food and pharma industries applications. This work followed the evolution of avocado fatty acids (FAs) and aliphatic acetogenins during seed germination and leaf growth. Oil extracts of embryonic axes, cotyledons, and leaves from seedlings and trees were divided to analyze free acetylated acetogenins (AcO-acetogenins), and free FAs. Embryonic axes from germinating seeds contained the highest amount of AcO-acetogenins and FAs; this tissue also accumulated the most diverse FA profile with up to 22 detected moieties. Leaves presented the highest variations in AcO-acetogenin profiles during development, although leaves from seedlings accumulated the simplest FA profile with only 10 different FAs. Remarkably, AcO-acetogenins represented half of the carbons allocated to lipids in grown leaves, while embryonic axes and cotyledons always contained more carbons within FAs during germination. Thus, we hypothesized the use of the AcO-acetogenin acyl chain for energy production toward β-oxidation. Also, α-linolenic and docosahexaenoic acids (DHAs) were proposed as close AcO-acetogenin intermediaries based on a correlation network generated using all these data. Another part of the oil extract was fractionated into different lipid classes before transesterification to profile FAs and acetogenins bound to lipids. Acetogenin backbones were identified for the first time in triglycerides from cotyledons and mainly in polar lipids (which include phospholipids) in all developing avocado tissues analyzed. Seed tissues accumulated preferentially polar lipids during germination, while triglycerides were consumed in cotyledons. Seedling leaves contained minute amounts of triglycerides, and polar lipids increased as they developed. Results from this work suggest acetogenins might be part of the energy and signaling metabolisms, and possibly of membrane structures, underlining the yet to establish role(s) of these unusual lipids in the avocado plant physiology.

Somatic embryogenesis and β-glucuronidase transformation in chickpea (Cicer arietinum cv. Bivanich).

Chickpea (Cicer arietinum L.) is an important kernel legume in the world. To optimize the plant tissue culture some experiments such as direct regeneration, proliferation, rooting shoots and somatic embryogenesis were done. In experiments were used direct regeneration and proliferation, various levels of plant growth regulators NAA (0 and 1.0mg/l), BAP (0, 1, 3 and 5mg/l) and three explants' types (epicotyl, cotyledon and embryonic axis). The results of both experiments showed that embryonic axis explant was better than other explants. The highest percentage was obtained in MS media containing 1mg/l BAP and also 3mg/l BAP and 0.1mg/l NAA with an average of 72%. The highest average number of branches (4.66) was found in the proliferation of embryonic axis in MS medium containing 3mg/l BAP. The highest rooting shoot (90%) was found in 1/2MS in B5 medium vitamins with 0.2mg/l of IBA and 0.5mg/l NAA. Somatic embryogenesis experiments were compared on the concentration gradient of 2,4-D in fine embryonic axis explants. The results displayed that the concentration gradient of 10mg/l 2,4-D to 5mg/l of 2,4-D and then to zero concentration showed the highest number of embryos. The best environment for regeneration embryos was MS medium with 2.5mg/l of 2,4-D concentration gradient to zero. In this study, the PCR reaction showed the presence of the β-glucuronidase (gus) marker gene in regenerated cotyledons for 20min in all three strains studied.

Does Cell Proliferation Affect Embryonic Flexure and Torsion?

Two vital and simultaneous morphogenetic events that occur during early embryo development are the flexure and torsion (F&T) of the embryonic body axis. Both contribute to development of the fetal position, which can be observed in avian and mammalian embryos. Correct progression of F&T is crucial for the proper development of the heart and other organs. Embryonic flexure can be divided into three segments: cranial, cervical, and thoracic. In this study, we focus on cervical flexure and torsion of the embryonic body axis. Differential cell proliferation, the difference in the rate of division of cells between regions in the developing embryo, is one mechanism that could influence F&T. Goodrum & Jacobson (1981) observed thickening of the dorsal midbrain relative to the ventral midbrain during cranial flexure. Miller & White (1998) have shown using mouse embryos that differential growth could cause torsion of an axis confined to an attached sphere.We performed studies inhibiting cell proliferation. Fertile white leghorn chick eggs were incubated to approximately Hamburger and Hamilton (HH) Stage 12 or 13. At these stages, F&T have just begun. We then added aphidicolin, a mitotic inhibitor, to whole embryo culture. Embryos were observed at 0, 5, and 10 hours to see if F&T were repressed. We hypothesized that the addition of aphidicolin would inhibit progression of F&T. We developed quantitative schemes to score the extent of F&T progression from HH 12 to HH 15. Embryos were scored from 0 to 4 based on the degree of cervical flexure and were also scored from 0 to 4 based on the degree of torsion. Afterward, the difference in F&T scores of same embryos at time t=0 h and t=10 h was calculated to determine the extent of both. These scores were then averaged for multiple embryos.Compared with controls (n=46), embryos incubated in 400 µM aphidicolin (n=18) flexed and rotated at a significantly lower rate. For flexion, the mean difference in scores for control and treated embryos were 1.85 and 1.22, respectively. For torsion, the mean difference in scores for control and treated embryos were 1.62 and 1.25, respectively. These differences were statistically significant. It is noted that global application of aphidicolin did impact the overall health of embryos. However, our preliminary results indicate that differential cell proliferation could play a role in rotation and flexure progression in the developing chicken embryo.

Preprint Highlight: Polarized Disheveled dissolution and condensation drives embryonic axis specification in oocytes.

Body axes establishment during animal embryogenesis often relies on the intrinsic asymmetry generated early on in the oocyte by the Wnt/β-catenin pathway. However, it's still unclear how the initial asymmetry arises. Here imaging experiments show that in starfish embryos, Disheveled (Dvl), an essential cytoplasmic transducer of Wnt signaling, becomes polarized at the vegetal pole via a dissolution-condensation mechanism coupled to meiotic progression. The authors further show that Dvl reformation is dependent on a maternally expressed Wnt signaling receptor Frizzled (Fzd1). This study identifies a novel link between the cell cycle and the body plan establishment and thus is relevant to a broad audience of developmental and evolutionary cell biologists.

Ultrastructural studies in Litchi (Litchi chinensis Sonn.) after desiccation and freezing

Recalcitrant seeds are remarkably short-lived and therefore are difficult to be successfully stored with respect to the ex situ conservation. Litchi seed embryonic axes were desiccated before cryopreserved in liquid nitrogen (direct immersion). Microscopic investigations were undertaken on Litchi embryonic axes in response to the desiccation and freezing sensitivity. Evidences shows that cryopreservation has a disruptive effect at the cellular level in Litchi.

Establishment of A Novel Plant Micropropagation System From Mature Hydroprimed Seeds Of Two Turkish Broad Bean Cultivars

Leguminous Broad bean (Vicia faba L.) is an excellent source of minerals, vitamins and proteins that makes it higly valuable food for human and feed for animal consumption. The plant has an important characteristic to fix atmospheric Nitrogen and play an important role to establish a natural balance of N in the atmosphere. It is a highly self-pollinated plant and has problem of low variation with limited genetic pool. Moreover, the previous studies identify problem of recalcitrance in broad bean. Therefore, there is need to establish a repeatable micropropagation protocol that could ensure an increase in genetic variability to overwhelm problems in breeding. This system must also be able for efficient gene delivery and could be integrated with the conventional breeding programs through direct organogenesis. The study aimed to develop a tissue culture protocol on two important Turkish broad bean cultivars Filiz99 and Eresen87 using MS medium containing 0.05, 0.15, 0.25, 0.35, 0.45, 0.55 mg l-1 TDZ using embryonic axis as explant. Maximum number of 5.33 and 3 shoots per explant were noted on cv. Filiz99 and Eresen87 on MS medium containing 0.15 mg l-1 TDZ. The developing shoots were rooted on MS medium containing 1 mg l-1 IAA after three weeks of culture. The rooted plants were transferred to pots containing peat under maintained under controlled greenhouse conditions for acclimatization The acclimatized plants bloomed and set seeds. Present results underscore importance of seed hydropriming before taking of explants to achieve high micropropagation on faba beans to overcome recalcitrance.

The nested embryonic dorsal domains of BMP-target genes are not scaled to size during the evolution of Drosophilaspecies.

Egg size is a fast-evolving trait among Drosophilids expected to change the spatial distribution of morphogens that pattern the embryonic axes. Here we asked whether the patterning of the dorsal region of the embryo by the Decapentaplegic/Bone Morphogenetic Protein-4 (DPP/BMP-4) gradient is scaled among Drosophila species with different egg sizes. This region specifies the extra-embryonic tissue amnioserosa and the ectoderm. We find that the entire dorsal region scales with embryo size, but the gene expression patterns regulated by DPP are not proportional, suggesting that the DPP gradient is differentially scaled during evolution. To further test whether the DPP gradient can scale or not in Drosophilamelanogaster, we created embryos with expanded dorsal regions that mimic changes in scale seen in other species and measured the resulting domains of DPP-target genes. We find that the proportions of these domains are not maintained, suggesting that the DPP gradient is unable to scale in the embryo. These and previous findings suggest that the embryonic dorso-ventral patterning lack scaling in the ventral and dorsal sides but is robust in the lateral region where the neuroectoderm is specified and two opposing gradients, Dorsal/NFkappa-B and DPP, intersect. We propose that the lack of scaling of the DPP gradient may contribute to changes in the size of the amnioserosa and the numbers of ectodermal cells with specific cortical tensions, which are expected to generate distinct mechanical forces for gastrulating embryos of different sizes.