Induction Of Embryogenesis Research Articles

SOMATIC EMBRYOGENESIS IN SAINFOIN (Onobrychis viciifolia Scop.)

Sainfoin (Onobrychis viciifolia Scop.) is a perennial legume used for forage production. The aim of this study was to investigate the effects of different basal media (Murashige&Skoog-MS, Schenk& Hildebrant-SH, Gamborg-B5 or CHU-N6), explant types (cotyledon and hypocotyl), and growth regulators (kinetin and thidiazuron) on the induction of somatic embryogenesis in sainfoin. For this purpose, hypocotyl and cotyledon explants isolated from in vitro grown sterile seedlings were cultured in solid MS, SH, B5, or N6 basal media supplemented with 2 mg/L 2,4-dichlorophenoxyacetic acid, 0.2 mg/L kinetin or thidiazuron, and 1 g/L proline. The developing embryonic callus was then transferred to a solid MS medium containing 3% sucrose. As a result of the experiment, callus weights per explant varied between 248.3-534.7 mg, 408.7-609.1, 220.3-537 mg and 127.3-428.3 mg in MS, B5, N6, and SH basal media respectively, depending on explants and growth regulators. Average callus weights were higher in B5 and N6 basal media than in MS and SH media, TDZ-containing media than kinetin containing media and cotyledon explant than hypocotyl explant. The highest frequency of explants developing somatic embryos (mean 33.3%) was obtained from kinetin-containing MS or B5 basal media, while the highest embryo per explant (mean 2.6 somatic embryo per explant) was obtained from kinetin-containing MS medium. The somatic embryo number per hypocotyl explant was higher than that of cotyledon explants.

Somatic Embryogenesis from the Leaf-Derived Calli of In Vitro Shoot-Regenerated Plantlets of Rosa hybrida 'Carola'.

Roses are one of the most important flowers applied to landscape, cut flowers, fragrance and food industries widely. As an effective method for plant reproduction, the regeneration via somatic embryos is the most promising method for breed improvement and genetic transformation of woody plants. However, lower somatic embryogenesis (SE) induction rates and genotypic constraints impede progress in genetic transformation in rose. This study describes a plant regeneration system for the famous red cut flower cultivar Rosa hybrida 'Carola'. The stems without petioles cultured on Murashige and Skoog (MS) medium supplemented with 1.0 mg·L-1 6-benzylaminopurine (6-BA), 0.05 mg·L-1 a-naphthalene acetic acid (NAA) and 30 g·L-1 sucrose showed the maximum proliferation coefficient of shoots with 3.41 for the micropropagation system. We evaluated the effects of different plant growth regulators (PGRs) on the induction, proliferation and conversion of somatic embryos. The induction rate of calli reached 100% on MS medium supplemented with 2.0 g·L-1 NAA and 30 g·L-1 glucose. The highest induction rate of somatic embryos achieved a frequency of 13.33% on MS medium supplemented with 2.0 mg·L-1 zeatin (ZT), 0.1 mg·L-1 NAA and 30 g·L-1 glucose. The most suitable carbohydrate with 60 g·L-1 glucose resulted in a proliferation rate of somatic embryos (4.02) on MS medium containing 1.5 mg·L-1 ZT, 0.2 mg·L-1 NAA and 0.1 mg·L-1 gibberellic acid (GA3). The highest somatic embryos germination rate (43.33%) was obtained from the MS medium supplemented with 1.0 mg·L-1 6-BA, 0.01 mg·L-1 IBA and 30 g·L-1 glucose. Finally, the germinated somatic embryos successfully rooted on 1/2 MS medium containing 1.0 mg·L-1 NAA, 30 g·L-1 sucrose, and the vigorous plantlets were obtained after hardening-off culture. This study provided a stable and efficient protocol for plant regeneration via somatic embryos in R. hybrida 'Carola', which will be beneficial to the further theoretical study and genetic improvement in roses.

Somatic Embryogenesis Induction in Coffea arabica L. by 2,4-Dichlorophenoxyacetic acid and 6-Furfurylaminopurine

Somatic Embryogenesis Induction in Coffea arabica L. by 2,4-Dichlorophenoxyacetic acid and 6-Furfurylaminopurine

Synergistic induction of fertilization-independent embryogenesis in rice egg cells by paternal-genome-expressed transcription factors.

In flowering plants, rapid activation of the zygotic genome occurs after fertilization1-3, but there is limited knowledge of the molecular pathways underlying embryo initiation4. In rice, a key role is played by the transcription factor BABY BOOM 1 (OsBBM1), initially expressed from the paternal genome1. Ectopic OsBBM1 expression in the egg cell can override the fertilization requirement, giving rise to parthenogenetic progeny5. Here we show that the WOX-family transcription factor DWARF TILLER1 (OsDWT1)/WUSCHEL-LIKE HOMEODOMAIN 9 (OsWOX9A)6, another gene paternally expressed in zygotes, is a strong enhancer of embryo initiation by OsBBM1. Co-expression of OsWOX9A and OsBBM1 in egg cells results in 86-91% parthenogenesis, representing 4- to 15-fold increases over OsBBM1 alone. These results suggest that embryo initiation is promoted by the synergistic action of paternal-genome-expressed transcription factors in the fertilized egg cell. These findings can be utilized for the efficient production of haploids, as well as clonal hybrid seeds in crop plants7,8.

Induction and maturation of somatic embryogenesis in mangaba tree a multipurpose Apocynaceae

This study reported the first results of in vitro somatic embryogenesis induction in Hancornia speciosa Gomes (mangaba tree, Apocynaceae). To induce callus, the TC accession leaf and nodal segments were cultivated in culture medium I (Murashige and Skoog; MS salts, putrescine, 45 µM 2,4-dichlorophenoxyacetic acid; 2,4-D and 22.19 µM 6-benzylaminopurine; BA). For the next stage (multiplication), the callus masses were sub-cultured into a culture medium II (MS salts, 22.62 µM 2,4-D and 22.19 µM BA). After 90 days, embryogenic calluses were inoculated to somatic embryo maturation medium culture III (MS salts, polyethylene glycol; PEG (0 and 2%) and BA (44.39 and 66.58 µM). Pro-embryos and somatic embryos in the maturation stage were assessed after 60 days. The rstatix package from R software with non-parametric tests (p < 0.05) were used to analysis data of experiments I and II. The Tukey’s test (p < 0.05) was performed on the data of experiment III. To induce and multiply embryogenic callus, 45 µM 2,4-D and 22.19 µM BA were efficient. More pro- and somatic embryo development were seen in the nodal explant cultured in the presence of 44.39 and 66.58 µM BA and 2% PEG. Putrescine does not promote the embryogenic masses of callus derived from leaf explants in the TC accession. Keywords: Hancornia speciosa Gomes, micropropagation, somatic embryo.

Analysis of the plant hormone expression profile during somatic embryogenesis induction in teak (Tectona grandis).

Plant somatic embryogenesis (SE) is an efficient regeneration system for propagation. It involves the regulation of a complex molecular regulatory network encompassing endogenous hormone synthesis, metabolism, and signal transduction processes, induced through exogenous plant growth regulators. Previous studies have focused primarily on traditional propagation methods for Tectona grandis, but there is limited knowledge on SE and its hormonal regulatory mechanisms. In our study, different SE stages, including the nonembryogenic callus (NEC), embryogenic callus (EC), and globular and heart-shaped embryo (E-SEs) stages, were induced in teak cotyledons incubated on MS medium supplemented with 0.1 mg/L thidiazuron (TDZ). Morphological and histological observations indicated that EC primarily originates from the development of embryogenic cell clusters. During SE induction, the levels of six classes of endogenous hormones, IAA, CTK, ETH, ABA, SA, and JA, changed significantly. Transcriptome analysis revealed that endogenous hormones participate in SE induction in teak through various biological processes, such as biosynthesis, metabolism, and signal transduction pathways. We found that IAA biosynthesis primarily occurs through the IAM pathway during these three stages. The ETH receptor kinase gene SERF1 exhibited the highest expression levels in E-SEs. The ABA-, SA-, and JA-related signal transduction genes ABI3, NPR1, and JAZ exhibited no differential expression during different stages. Moreover, key encoding genes of SE regulators, including WUS, WOX and SERK, were differentially expressed during SE. In conclusion, this study offers insights into the roles of endogenous hormones and their interactions during SE induction.

Endogenous phytohormone analysis during sequential stages of somatic embryogenesis in Musa AAA cv. Grand Naine (subgroup Cavendish)

Abstract This is the first initiative study to provide quantified levels of four endogenous phytohormones during the induction, regeneration, maturation and germination stages of somatic embryogenesis (SE) in banana, using immature male bud explants, in the popular commercial triploid cultivar Grand Naine (AAA). Phytohormones are known to play a major role in the regulation of SE and most protocols have been developed for various cultivars of banana following empirical approaches. Although, several reports are available with regard to the impact of growth regulators exogenously supplemented to the media, the vital role of endogenous growth hormones in various developmental stages of SE remain unclear in banana. Reverse Phase High Performance Liquid Chromatography (RP-HPLC) was used to analyze endogenous status of indole acetic acid (IAA), zeatin, gibberellic acid (GA) and abscisic acid (ABA). Results from this study revealed that endogenous IAA and GA along with exogenous auxin play a major role in the induction of embryogenic competence of male bud explants. The levels of endogenous IAA and zeatin were found to be higher in embryogenic calli than non-embryogenic calli, although three different types of exogenous auxins were supplemented in callogenesis medium, but not cytokinin. Higher level of ABA and GA was observed in early and late developmental stages of somatic embryo, respectively. GA also played a predominant role in germination of somatic embryos. These findings will be applied for improvement of SE protocol in recalcitrant banana cultivars by suitably altering or supplementing exogenous growth hormones.

MiR156-SPL and miR169-NF-YA Modules Regulate the Induction of Somatic Embryogenesis in Arabidopsis via LEC- and Auxin-Related Pathways.

The embryogenic transition of plant somatic cells to produce somatic embryos requires extensive reprogramming of the cell transcriptome. The prominent role of transcription factors (TFs) and miRNAs in controlling somatic embryogenesis (SE) induction in plants was documented. The profiling of MIRNA expression in the embryogenic culture of Arabidopsis implied the contribution of the miR156 and miR169 to the embryogenic induction. In the present study, the function of miR156 and miR169 and the candidate targets, SPL and NF-YA genes, were investigated in Arabidopsis SE. The results showed that misexpression of MIRNA156 and candidate SPL target genes (SPL2, 3, 4, 5, 9, 10, 11, 13, 15) negatively affected the embryogenic potential of transgenic explants, suggesting that specific fine-tuning of the miR156 and target genes expression levels seems essential for efficient SE induction. The results revealed that SPL11 under the control of miR156 might contribute to SE induction by regulating the master regulators of SE, the LEC (LEAFY COTYLEDON) genes (LEC1, LEC2, FUS3). Moreover, the role of miR169 and its candidate NF-YA targets in SE induction was demonstrated. The results showed that several miR169 targets, including NF-YA1, 3, 5, 8, and 10, positively regulated SE. We found, that miR169 via NF-YA5 seems to modulate the expression of a master SE regulator LEC1/NF-YA and other auxin-related genes: YUCCA (YUC4, 10) and PIN1 in SE induction. The study provided new insights into miR156-SPL and miR169-NF-YA functions in the auxin-related and LEC-controlled regulatory network of SE.

Induction of Somatic Embryogenesis of Pontianak Siamese Orange Cotyledon Cultures on Murashige Skoog Media with the Addition of 2,4-D and Kinetin

Conventional citrus propagation has weaknesses, one of which is susceptible to CVPD(Citrus Vein Phloem Degeneration) and other diseases which are the main cause of the decline in population and production of pontianak siam orange plants in West Kalimantan. One alternative to in vitro propagation of citrus seedlings is the induction of somatic embryogenesis. The purpose of the study was to obtain the best combination of 2,4-D and kinetin concentrations for callus induction and somatic embryo induction from Pontianak siam orange cotyledon explants, observing the developmental phase (stage) of embryos formed. The implementation of the study used a factorial completely randomized design (CRD) consisting of 2 factors. The first factor is the concentration of 2,4-D consisting of 4 concentration levels, namely 0; 0.5; 1; 1.5; 2 mg/L. The second factor is kinetin concentration consisting of 4 concentration levels, namely 0; 0.5; 1; 1.5; 2 mg/L.The observation parameters include the time of callus emergence (hst), the percentage of explants forming callus (%), the texture and color of the callus, observing the stage of the embryo found. The results showed that 72% of treatments were able to induce callus from pontianak siam orange cotyledons with an average callus appearance at 7 hst. The callus formed is compact and crumbly with varied callus colors, namely white, yellowish, greenish and brownish. The best treatment that produces embryogenic callus is the combination of D0.5K0.5; D1K1; D0.5K1.5; D1.5K1.5 and D2K2 with the characteristics of crumbly callus and yellowish and brownish colors. The results of microscopic observations found that the somatic embryo phase formed was the pro-embryo phase.

Introduction of Salicornia europaea L into in vitro culture

The goal of the research was to develop a technology for introducing Salicornia europaea L.into in vitro culture. Methods of cultivating meristems and callus cultures were studied. To cultivate meristems, the tip of the apical bud were used as an explant. Fragments of stems and leaves were used toobtain callus tissue. To study the influence of various factors on germination, seeds were soaked in sterile tap water and in solutions of gibberellin, cytokinin, auxin and NaCl, and were also subjected to cold stratification (independently and with subsequent placement in Knop’s agarized medium). Salicornia seeds were sterilized with various antiseptics: 70% alcohol, 10% aqueous solution of sodium hypochlorite («Belizna»), amoxicillinand 3% hydrogen peroxide. The ability of the culture to form callus was studied in MS medium. As a result, it was determined that the highest germination of seeds was observed in Knop medium after treating theseeds with a suspension of green algae of the Scenedesmus genus, as well as after preliminary cold stratification, and slightly less after treating the seeds with a solution of NaCl and gibberellic acid. The most effective method of seed sterilization turned out to be treatment with alcohol followed by treatment with sodium hypochlorite. A comparative analysis of seed germination in filter paper in Petri dishes, Knop agar medium, Murashige and Skoog (hormone-free) was carried out. The ability of S. europaea L. to form callusin MS medium with phytohormones was assessed. Conclusion. To improve germination, it is recommended to subject the seeds to cold stratification. To obtain aseptic explants quickly, it is recommended to germinate the seeds in the Knop nutrient medium, having previously sterilized them with alcohol, then with sodiumhypochlorite, followed by washing with distilled water. The most suitable type of microclonal propagationfor S. europaea L. is the production of callus tissue followed by induction of organogenesis or embryogenesis.

Synergistic effects of L-glutamine and inorganic nitrogen molar ratios enhance the induction of somatic embryogenesis of Pinus maximinoi H.E. Moore

Clonal breeding programs of Pinus maximinoi require the establishment of a robust somatic embryogenesis (SE) protocol to produce enough cell lines to accelerate the effective continuous deployment of elite planting stocks to research and commercial compartments. Somatic embryogenesis was induced from immature zygotic embryo explants enclosed in megagametophytes of P. maximinoi collected from two plantations located in different climatic conditions. Cones were collected during the winter months from July to August and the influence of seed family, cone collection date and culture medium formulation, with emphasis on the organic and inorganic nitrogen supply, were studied. Ammonium to nitrate molar ratios of 1:1 and 1:2 in modified Litvay’s medium (mLV) produced the highest numbers of extrusions, while a 1:4 ratio mostly produced unhealthy, non-embryogenic extrusions. The formation of a tissue showing a rapidly-proliferating, spiky morphotype was produced in a medium supplemented with 1.5 g/L of L-glutamine. Morphologically advanced cultures with nodular structures were produced in megagametophytes from both plantations in a 1:2 NH4+:NO3− medium regardless of L-glutamine supplementation levels. The optimal medium for P. maximinoi SE induction contained a 1:2 NH4+:NO3− molar ratio with 1.5 g/L L-glutamine. The synergy between the molar ratio of NH4+:NO3− and L-glutamine resulted in the highest numbers of extrusions. The overall inductive competence window for somatic embryogenic response in P. maximinoi was determined to be from the second week of July to the first week of August for both plantations. The “peak” period was in the fourth week of July 2022. The success of the SE technology in P. maximinoi seed families is determined by the optimal inductive competence window of the immature megagametophytes enclosing zygotic embryos and the chemical composition of the induction medium in terms of the ammonium to nitrate molar ratio and the concentration of the L-glutamine used.

Whole-transcriptome sequence analysis of pumpkin ovules in response to heat shock during unfertilized ovary culture

Unfertilized ovary culture is one of the most effective approaches to induce haploids in Cucurbitaceae. Previous studies have greatly contributed to inducing more and better haploids, but there has been less contribution to the mechanism involved. Our previous research established efficient haploid regeneration protocol of unfertilized ovary culture and found that heat shock can effectively promote ELSs’ induction. In this study, firstly, we accurately calculated the contribution of heat shock to ELSs’ induction via unfertilized ovary culture of pumpkin, and then performed whole-transcriptome sequence using “Yi-Chuan-Ling 4″, which is more sensitive to heat shock and has a higher induction rate. The results show that heat shock treatment was obviously beneficial to the unfertilized ovary culture of pumpkin, and the average induction rate in the treatment group (35°C,5d) was 26.6 times higher than in the control group (25°C,5d) . Through whole-transcriptome sequencing, we obtained 4579 differentially expressed (DE) RNAs, including 3133 mRNAs, 120 lncRNAs, 814 circRNAs, and 512 miRNAs. GO annotation analysis showed that DEmRNAs were associated with cellular processes, metabolic processes, biological regulation, response to stimulus, and so on. The ceRNA network was constructed using differential expressed RNAs. It captured the gene MSTRG.28850.1, which was related to embryogenesis and regulated by miRNA Nov-m0336–5 p. In conclusion, these findings might provide an important reference for mechanism research regarding embryogenesis induction and accelerate the application of unfertilized ovary/ovule culture in Cucurbita breeding.

New Prospects for Improving Microspore Embryogenesis Induction in Highly Recalcitrant Winter Wheat Lines

Among various methods stimulating biological progress, double haploid (DH) technology, which utilizes the process of microspore embryogenesis (ME), is potentially the most effective. However, the process depends on complex interactions between many genetic, physiological and environmental variables, and in many cases, e.g., winter wheat, does not operate with the efficiency required for commercial use. Stress associated with low-temperature treatment, isolation and transfer to in vitro culture has been shown to disturb redox homeostasis and generate relatively high levels of reactive oxygen species (ROS), affecting microspore vitality. The aim of this study was to investigate whether controlled plant growth, specific tiller pre-treatment and culture conditions could improve the potential of microspores to cope with stress and effectively induce ME. To understand the mechanism of the stress response, hydrogen peroxide levels, total activity and the content of the most important low-molecular-weight antioxidants (glutathione and ascorbate), as well as the content of selected macro- (Mg, Ca, NA, K) and micronutrients (Mn, Zn, Fe, Cu, Mo) were determined. These analyses, combined with the cytological characteristics of the microspore suspensions, allowed us to demonstrate that an increased microspore vitality and stronger response to ME induction were associated with higher stress resistance based on more efficient ROS scavenging and nutrient management. It was shown that a modified procedure, combining a low temperature with mannitol and sodium selenate tiller pre-treatment, reduced oxidative stress and improved the effectiveness of ME in winter wheat lines.

Study on the induction of somatic embryogenesis and morphological structural changes during the development of Handeliodendron bodinieri L.

Study on the induction of somatic embryogenesis and morphological structural changes during the development of Handeliodendron bodinieri L.

Transcriptomic Analysis During the Induction of Somatic Embryogenesis in Coffea canephora.

Omic tools have changed the way of doing research in experimental biology. The somatic embryogenesis (SE) study has not been immune to this benefit. The transcriptomic tools have been used to compare the genes expressed during the induction of SE with the genes expressed in zygotic embryogenesis or to compare the development of the different stages embryos go through. It has also been used to compare the expression of genes during the development of calli from which SE is induced, as well as many other applications. The protocol described here is employed in our laboratory to extract RNA and generate several transcriptomes for the study of SE on Coffea canephora.

Proteomic Approach during the Induction of Somatic Embryogenesis in Coffea canephora.

Plant growth regulators (PGR) are essential for somatic embryogenesis (SE) in different species, and Coffea canephora is no exception. In our study model, previously, we have been able to elucidate the participation of various genes involved in SE by using different strategies; however, until now, we have not used a proteomic approach. This research seeks to contribute to understanding the primary cellular pathways involved in developing SE in C. canephora. The process of our model consists of two stages: (1) preconditioning in MS medium with auxin (NAA) and cytokinin (KIN), and (2) induction in Yasuda liquid medium added with cytokinin (BA). Therefore, in this study, we analyzed different days of the SE induction process using shotgun label-free proteomics. An amount of 1630 proteins was found among different sampling days of the process, of which the majority were accumulated during the induction stage. We found that some of the most enriched pathways during this process were the biosynthesis of amino acids and secondary metabolites. Eighteen proteins were found related to auxin homeostasis and two to cytokinin metabolism, such as ABC, BIG, ILR, LOG, and ARR. Ten proteins and transcription factors related to SE were also identified, like SERK1, SKP1, nuclear transcription factor Y, MADS-box, and calreticulin, and 19 related to other processes of plant development, among which the 14-3-3 and PP2A proteins stand out. This is the first report on the proteomic approach to elucidate the mechanisms that operate during the induction of SE in C. canephora. So, our findings provide the groundwork for future, more in-depth research. Data are available via ProteomeXchange Consortium via the PRIDE partner repository with the dataset identifier PXD047172.

The of Effects of Light and a Combination of Growth Regulators on the Induction of Somatic Embryogenesis in Orchid Rhynchostylis gigantea (LindI.) Ridl.

The squirrel-tail orchid (Rhynchostylis gigantea) belongs to the Orchidaceae family. This orchid is indigenous to Southeast Asia and is scented and arranged in a bouquet of dangling stems. Its uniqueness makes it commercially desirable. In vitro culture has been utilized for orchid proliferation for a very long time to aid in the development of orchid seedlings. However, not all orchid species react the same way. Through somatic embryogenesis without gamete fusion, in vitro culture techniques can produce new plants during the entire embryonic phase. Endogenous and exogenous hormones, as well as light, influence the success of embryogenesis. Determining the optimal environmental parameters (light, combination and concentration of growth regulators, and their interaction) for inducing somatic embryogenesis from leaf explants in R. gigantea is the purpose of the present study. The leaves of R. gigantea clone 19 were utilized as explants in this investigation, utilizing a completely random factorial design. The first factor is a combination of growth regulator types and concentrations, and the second factor is light. Explants that are alive and growing at a rapid rate indicate that the in vitro culture is successful. Light and the combination of growth regulators significantly affected the percentage of viable explants, the beginning date of callus formation, and the number of embryogenic calluses produced. The interaction of two components (light and a combination of growth regulators) did not affect the three characteristics, with the exception of the proportion of somatic embryogenesis. Incubation in the dark is the optimal environment for initiating somatic embryogenesis in explants. The optimal combination of plant growth regulators for inducing somatic embryogenesis was 0.5 mgL-1 TDZ and 0.1 mgL-1 BAP. Bright light and a concentration of 1.0 mgL-1 TDZ + 0.1 mgL-1 BAP were the optimal interaction conditions for the induction of somatic embryogenesis in R. gigantea.

Transcriptomics integrated with metabolomics reveals the effect of cold stress on rice microspores

BackgroundMicrospore culture is one of the important biotechnological tools in plant breeding. The induction of microspore embryogenesis is a critical factor that affects the yield of microspore-derived embryo productions. Cold treatment has been reported to reprogram the gametophytic pathway in various plant species. However, the exact mechanism(s) underlying the effect of cold pre-treatment of floral buds on the efficiency of ME is still not clear.ResultsIn this study, the effects of cold stress on the microspore totipotency of rice cultivar Zhonghua 11 were investigated. Our results revealed that a 10-day cold treatment is necessary for microspore embryogenesis initiation. During this period, the survival rate of microspores increased and reached a peak at 7 days post treatment (dpt), before decreasing at 10 dpt. RNA-seq analysis showed that the number of DEGs increased from 3 dpt to 10 dpt, with more downregulated DEGs than upregulated ones at the same time point. GO enrichment analysis showed a shift from ‘Response to abiotic stimulus’ at 3 dpt to ‘Metabolic process’ at 7 and 10 dpt, with the most significant category in the cellular component being ‘cell wall’. KEGG analysis of the pathways revealed changes during cold treatment. Mass spectrometry was used to evaluate the variations in metabolites at 10 dpt compared to 0 dpt, with more downregulated DEMs being determined in both GC-MS and LC-MS modes. These DEMs were classified into 11 categories, Most of the DEMs belonged to ‘lipids and lipid-like molecules’. KEGG analysis of DEMs indicates pathways related to amino acid and nucleotide metabolism being upregulated and those related to carbohydrate metabolism being downregulated. An integration analysis of transcriptomics and metabolomics showed that most pathways belonged to ‘Amino acid metabolism’ and ‘Carbohydrate metabolism’. Four DEMs were identified in the interaction network, with stearidonic acid involving in the most correlations, suggesting the potential role in microspore totipotency.ConclusionsOur findings exhibited the molecular events occurring during stress-induced rice microspore. Pathways related to ‘Amino acid metabolism’ and ‘Carbohydrate metabolism’ may play important roles in rice microspore totipotency. Stearidonic acid was identified, which may participate in the initiation of microspore embryogenesis.

Plant regeneration through somatic embryogenesis in cell suspensions of Cenchrus ciliaris L.

BackgroundThis study aims to establish cell suspension culture, its maintenance and induction of somatic embryogenesis, and in vitro plant regeneration in Cenchrus ciliaris L. Suspension cultures are relatively homogenous cell lines facilitating uniform access to nutrition. These are ideal sources of competent cells for genetic transformation.ResultsCallus was initiated by culturing immature inflorescences of Cenchrus ciliaris cv. IGFRI-3108 on Murashige and Skoog (MS) medium containing 3 mg l−1 2,4-dichlorophenoxyacetic acid (2,4-D) and 0.5 mg l−1 6-benzylaminopurine (BAP) with 30 g l−1 sucrose. Cell suspension cultures were established in liquid MS medium using an inoculum size of 10 g l−1. These were maintained to achieve embryogenic cell/regeneration competent cultures. Growth curve analysis and a subculture interval of 20 days were determined to harvest cells at the end of the exponential phase. The cell doubling time was found to be 11 days. Somatic embryogenesis was accomplished in MS medium containing 1 mg l−1 2,4-D, 1 mg l−1 BAP along with growth adjuvants, 300 mg l−1 casein hydrolysate, 400 mg l−1 glutamine and 300 mg l−1 proline. The highest number (16 ± 3.78/per inoculum) of shoots regenerated on this medium. The elongation and rooting of shoots were recorded on basal MS and ½ MS media, respectively. Rooted plants were successfully transferred to pots containing a Soilrite and cocopeat mixture in a 3:1 proportion for 3–4 weeks and later successfully acclimatized in the greenhouse with a 60% survival rate. The genetic fidelity of 12 regenerated plants was analysed using RAPD primers that were genetically identical to the mother plant.ConclusionCell suspension culture-based in vitro plant regeneration of C. ciliaris involved the establishment, maintenance and progression of somatic embryogenesis during shoot and root development. The inherent limitation of callus-mediated in vitro plant regeneration reducing the regeneration potential due to the aging of the calli has been overcome.

Pioneering anther culture-based embryogenesis in Solanum aethiopicum L.

This study aimed to establish a compelling approach for inducing embryogenesis through in vitro anther culture in Scarlet eggplant (Solanum aethiopicum L.), the most consumed and popular eggplant among indigenous vegetables on the African continent. While in vitro androgenesis has been favorably employed in brinjal (Solanum melongena L.) breeding, there has been no attempt to induce embryogenesis in a large germplasm of its relative, Solanum aethiopicum. In two distinct experiments, the largest germplasm collection of Solanum aethiopicum gr. Gilo was assessed for embryogenesis induction using C medium supplemented with different concentrations of hormones. In the first experiment, callus induction was successful with an overall rate of 36.6 calli/100 anthers, but embryo formation was unsuccessful. Statistical analysis revealed a dependency of the rate of callus induction on accessions. In the second experiment, only four selected accessions of Solanum aethiopicum gr. Gilo were used and compared to two Turkish eggplant genotypes of Solanum melongena in two distinct treatments. The results showed that in the first treatment (I), only the accession GKE12 had a satisfactory outcome with a rate of embryo formation of 0.82/100 anthers and 0.41/100 anthers corresponding to the rate of developed embryos. In the second treatment (II), only controls, which were Adana and Kemer cultivars of Solanum melongena formed embryos with a rate of 7.26/100 anthers and 1.15/100 anthers, respectively. The obtained embryo/seedling of Solanum aethiopicum gr. Gilo was found to be diploid. Overall, this study demonstrated that with the right combinations of hormones, it is possible to induce embryogenesis and produce a diploid of Solanum aethiopicum, the world’s second most popular cultivated eggplant after brinjal. These findings could potentially contribute to the breeding of eggplants for enhanced genetic variation and resistance.