μM ABA Research Articles

Screening and Identification of Target Gene of StTCP7 Transcription Factor in Potato

TCP transcription factors are involved in the regulation of plant growth and development and response to stress. Previous studies showed that StTCP7 was involved in the abiotic stress response of potato and positively regulated plant tolerance to drought stress. On the basis of previous studies, this study verified the downstream target genes of StTCP7 transcription factor binding through yeast one hybridization, double luciferase and other technologies, and conducted a preliminary analysis of the downstream target genes. The results showed that the StTCP7 transcription factor could bind the promoter region of StDAM5 and StGOLS2 and regulate the expression of their genes. qRT-PCR analysis showed that the expression level of StDAM5 gene was the highest in flower stalk tissue and the lowest in leaf stalk. The expression of StGOLS2 gene was the highest in stem, the second in stalk, and the lower in root. Both StDAM5 and StGOLS2 genes responded to abiotic stress treated with 200 mM NaCl, 20% PEG-6000 and 100 µM ABA. The expression levels of target genes StDAM5 and StGOLS2 were up-regulated in StTCP7 interfered plants. The protein encoded by the target gene StDAM5 belongs to the Type II MADS-box protein, which contains 238 amino acids and is an acidic hydrophilic protein. The analysis of StDAM5 promoter region showed that the promoter region of StDAM5 gene contained cis-acting elements such as light response and abscisic acid. Subcellular localization showed that StDAM5 protein was expressed in both nucleus and cytoplasm.

Maize Autophagy-Related Protein ZmATG3 Confers Tolerance to Multiple Abiotic Stresses.

Abiotic stresses pose a major increasing problem for the cultivation of maize. Autophagy plays a vital role in recycling and re-utilizing nutrients and adapting to stress. However, the role of autophagy in the response to abiotic stress in maize has not yet been investigated. Here, ZmATG3, which is essential for ATG8-PE conjugation, was isolated from the maize inbred line B73. The ATG3 sequence was conserved, including the C-terminal domains with HPC and FLKF motifs and the catalytic domain in different species. The promoter of the ZmATG3 gene contained a number of elements involved in responses to environmental stresses or hormones. Heterologous expression of ZmATG3 in yeast promoted the growth of strain under salt, mannitol, and low-nitrogen stress. The expression of ZmATG3 could be altered by various types of abiotic stress (200 mM NaCl, 200 mM mannitol, low N) and exogenous hormones (500 µM ABA). GUS staining analysis of ZmATG3-GUS transgenic Arabidopsis revealed that GUS gene activity increased after abiotic treatment. ZmATG3-overexpressing Arabidopsis plants had higher osmotic and salinity stress tolerance than wild-type plants. Overexpression of ZmATG3 up-regulated the expression of other AtATGs (AtATG3, AtATG5, and AtATG8b) under NaCl, mannitol and LN stress. These findings demonstrate that overexpression of ZmATG3 can improve tolerance to multiple abiotic stresses.

Unveiling machine learning's impact on in vitro callogenesis optimization in Crocus sativus L

Saffron is a spice derived from its flower (Crocus sativus L.) and it is grown for its medicinal and culinary uses worldwide. Callus is an intermediate stage between explant and in vitro organogenesis. The fate of embryogenesis and organogenesis depends on the quality of the callus. Callogenesis is a complex biological process and is affected by various intrinsic and extrinsic factors. Machine learning (ML), can effectively cope with the time and cost constraints of experiments. In this study, we emphasized the effects of sucrose and phytohormones (auxin, and abscisic acid) on callogenesis with the aid of a machine-learning algorithm. Among the different tested hormone combinations, maximum callus proliferation was observed in the MS media plate containing 3 % sucrose, supplemented with 100 µM ABA, 2 and 4 mg/L NAA, and 2,4-D. A total of eight different ML tools were used to predict the efficacy of inputs on callogenesis. Out of all applied algorithms, Xtreme Gradient Boosting (XGB) and Gradient Boosted Regression (GBR) provide better prediction performance than other models on callus induction and proliferation. SHAP analysis reveals the significance of sucrose, plant growth regulators (NAA, 2,4-D, and ABA), and time on callogenesis. Therefore, machine learning programs can effectively save cost and time by guiding experiments and facilitating optimization.

OsAAI1 Increases Rice Yield and Drought Tolerance Dependent on ABA-Mediated Regulatory and ROS Scavenging Pathway

In this study, we investigated the function of OsAAI1 in yield and drought tolerance by constructing overexpression line OE-OsAAI1 and mutant line osaai1. Bioinformatics analysis showed that the AAI gene-OsAAI1- belongs to the HPS_like subfamily of the AAI_LTSS superfamily, and OsAAI1 was localized in the nucleus. The expression of OsAAI1 was significantly induced by ABA and drought stress. OsAAI1 overexpression (OE19) significantly increased, and gene mutant (osaai1-1) repressed plant height, primary root length, lateral root number, grain size and yield in rice. Moreover, physiological and biochemical analyses showed that osaai1 was sensitive to drought stress, while OE19 enhanced the drought tolerance in rice. DAB and NBT staining revealed that under drought treatment, osaai1 accumulated a large amount of ROS compared with the wild type, while OE19 accumulated the least, and CAT, APX, GPX, GR activities were higher in OE19 and lower in osaai1, suggesting that OE19 improves rice tolerance to drought stress by enhancing ROS scavenging ability. OE19 also induce the expression of ABA-mediated regulatory pathway genes and enhance accumulation of ABA content in rice seedling. Predictably, OE19 displayed enhanced sensitivity to ABA, and ROS accumulation was significantly higher than in wild type and osaai1 under 3 µM ABA treatment. Thus, these results suggest that OsAAI1 is a positive regulator of rice yield and drought tolerance dependent on the ABA-mediated regulatory and ROS scavenging pathway.

Ethylene Signaling Pathway Genes in Strawberry and Their Expression Patterns during Fruit Ripening

Ethylene at least partly regulates some aspects during non-climacteric ripening in strawberry. However, the ethylene signaling pathway genes in the strawberry fruit have not been comprehensively and systematically analyzed. In the present study, 15 FaETRs and 14 FaEIN3/EINs were identified in the octoploid strawberry genome. Subcellular localization analysis predicted that FaETRs and FaEIN3/EINs are respectively localized to the endoplasmic reticulum and the nucleus. The phylogenetic trees showed that FaETRs were classified into two subgroups, while FaEIN3/EINs were divided into three clades, which was supported by gene structure and conserved motif analysis. FaETRs and FaEIN3/EINs could interact with several components, such as CTR1, RTE1, EIN2 and ERF1B, in the ethylene signaling pathway by protein–protein interaction network analysis. Transcriptomic data showed that FaETRs were mainly expressed at the early stage of fruit development in three strawberry cultivars. Additionally, a couple of FaETRs (FaETR2 and FaETR13) and FaEINs (FaEIN2 and FaEIN7) could be induced by 1 μM ABA and inhibited by 100 μM nordihydroguaiaretic acid (NDGA, an ABA biosynthesis blocker). These findings suggested that the FaETR- and FaEIN3/EIN-mediated ethylene signaling pathway might play a role in strawberry fruit ripening.

Self-pruning in lime (<i>Citrus aurantifolia</i> Swingle) after treatments with ichiphon, abscisic acid and nitrogen, phosphorus, and potassium fertilizers

Self-pruning can lower production costs, especially in the case of annual crops such as citrus. The study aimed to determine self-pruning of lime treated with growth regulators and a fertilizer. Self-pruning was applied on a one-year-old of <i>Citrus aurantifolia</i> from February 2020 to April 2022 in Bogor, Indonesia. The treatments used NPK at three levels: 22.5:7.5:2.5 g/tree, 32.5:17.5:12.5 g/tree, and 42.5:27.5:22.5 g/tree, and growth regulators: 500 ppm ichiphon + 100 µM ABA, 750 ppm ichiphon + 50 µM ABA, and 1000 ppm ichiphon. Initial symptoms of self-pruning, namely leaf fall, which correlated with the ethylene concentration in the leaves, occurred in all treatment applications. The combination of the NPK fertilizer 32.5:17.5:12.5 g/tree with 750 ppm ichiphon + 50 M ABA gave the highest ethylene concentration. The highest concentration of ABA was found on the first day after the treatment with the NPK fertilizer 42.5: 27:5: 22.5 g/tree and growth regulators 500 ppm ichiphon + 100 M ABA; however, it did not differ from the treatment with the NPK fertilizer 42.5: 27.5:22.5 g/tree and the 1000 ppm ichiphon growth regulator on the fourth and twelfth days. The percentage of secondary, tertiary, and deciduous branches did not differ between the treatments. Self-pruning that occurs as a result of induction by ichiphon, abscisic acid, and NPK fertilizers, can be an alternative to mechanical pruning.

The synergistic effect of ascorbic and abscisic acids on enriching AVC-dominated avenanthramides in oat germination process

Avenanthramides are a group of antioxidants, of which avenanthramide C (AVC) shows the strongest antioxidant activity. Germination is conducive to the accumulation of total avenanthramides. However, it is unclear how to regulate the proportion of AVC in total avenanthramides by controlling the oat-germination process. To increase the proportion of AVC in total avenanthramides, single or combined treatments with abscisic (ABA) and ascorbic acid (Asc) during the steeping or germination period was performed. Compared to the control (541 μg/g dry weight), the 300 μM-ABA treatment during the steeping period significantly increased the proportion of AVC in total avenanthramides from 30.93 to 40.70%, which was accompanied by a remarkable increase in the content of total avenanthramides (1.8- fold). The content of total avenanthramides and the proportion of AVC within increased significantly (3.35-fold and by 9.28%, respectively) compared to the control when successively treated with 300 μM ABA during the steeping period and 20 mM Asc during the germination period. Moreover, ABA and Asc were synergistic in increasing the activities of hydroxycinnamoyl-CoA:hydroxyanthranilate N-hydroxycinnamoyl transferase (HHT) and tyrosine ammonia-lyase (TAL), but reduced the activity of peroxidase (POD), which resulted in the accumulation of AVC-dominated avenanthramides. These findings provide novel information on the bio-concentration of AVC by regulation of the oat-germination process.

Overexpression of a C3HC4-type RING E3 ligase gene, OsRFPHC-13, improves salinity resistance in rice, Oryza sativa, by altering the expression of Na+/K+ transporter genes

Salinity stress is a major restriction to increasing crop production. We characterized the Oryza sativa C3HC4-type RING E3 ligase-13 (OsRFPHC-13), which contributes a positive role in enhancing salinity insensitivity. Expression of OsRFPHC-13 was highly induced by treatment with 100 mM NaCl, 20% PEG, or 100 µM ABA. The OsRFPHC-13-GFP fusion protein was expressed in the chloroplasts and cytosol of the rice protoplasts. An in vitro ubiquitination assay confirmed that OsRFPHC-13 possesses E3 ligase activity; however, the mutation of OsRFPHC-13C232A did not show this activity. OsRFPHC-13-overexpressing plants showed a dwarf phenotype, short primary root length, and small lateral root number compared to WT and osrpfhc-13. In contrast, under salt stress, OsRFPHC-13-overexpressing plants showed enhanced salinity insensitivity by reducing the accumulation of Na+ in shoots and roots, increasing the activity of ROS scavenging, and positively regulating the expression of Na+/K+ transporter genes compared to WT and osrpfhc-13. In addition, OsRFPHC-13-overexpressing plants exhibited increased ABA sensitivity in the germination assay but decreased sensitivity in osrpfhc-13 compared to WT. These results suggest that OsRFPH-13 E3 ligase positively regulates the response to salinity via an ABA-dependent mechanism in rice.

The identification of suitable internal reference genes in quinoa seeds subjected to abscisic acid and gibberellin treatment

Abstract: Quinoa has been recognized as the sole “comprehensive nutritional crop”; however, it is susceptible to pre-harvest sprouting (PHS). While quantitative reverse transcription polymerase chain reaction (RT-qPCR) has been extensively employed for gene expression level detection, the selection of suitable reference genes is imperative to ensure precise gene expression quantification across diverse conditions. This study aims to identify stable reference genes in quinoa seeds under ABA and GA, in order to provide a basis for subsequent research on PHS. Seeds were subjected to different concentrations of ABA and GA (10 μM, 50 μM, 100 μM, and 200 μM). The most suitable treatment concentration was determined based on seed viability. Here, MON1, GAPDH, EIF3, EF1α, ACT, TUB1, and TUB6 were selected as candidate genes. The suitability of these reference genes under different conditions was assessed using various methods including Ct values, geNorm, NormFinder, BestKeeper, Delta Ct, and RefFinder. Based on the results obtained from the hormone experiments, it was observed that the application of 100 μM ABA and 200 μM GA yielded the most advantageous outcomes. Additionally, the most appropriate reference genes for different treatments are ACT and TUB1 (H2O treatment), EIF3 and MON1 (ABA, GA treatment and also for the combined data set of the three groups). However, GAPDH exhibited the least stability across all treatments. In summary, ACT is recommended as the reference gene for natural quinoa germination, while EIF3 and MON1 should be used for ABA and GA treatments.

Abscisic Acid Improves Linoleic Acid Accumulation Possibly by Promoting Expression of EgFAD2 and Other Fatty Acid Biosynthesis Genes in Oil Palm Mesocarp.

Abscisic acid plays an important role in fruit development. However, the effect of ABA on fatty acid biosynthesis in oil palm is still unknown. In this study, ABA treatments (CK, A1–A4) were applied to oil palm fruit at 16 WAP (weeks after pollination), and fatty acids in the mesocarp at 24 WAP were analyzed by GC-MS. Results showed that linoleic acid content under treatment A2 (20 μM ABA) was significantly higher (slightly increased by 8.33%) than the control. Therefore, mesocarp samples of A2, and the control at 16, 20, and 24 WAP was sampled for RNA-Seq. KEGG pathway enrichment analysis showed that 43 genes were differentially expressed in the fatty acid biosynthesis pathway, of which expression of EgFAD2 (unigene 105050201) under 20 μM ABA treatment was 1.84-fold higher than in the control at 20 WAP. Further sequence analysis found that unigene 105050201 had more ABA-responsive elements (ABRE), complete conserved domains, and a C-terminal signaling motif among two FAD2 copies. Furthermore, WGCNA and correlation analysis showed co-expression of EgFAD2 (unigene 105050201) with transcription factors (TFs) (WRI1, AP2-EREBP, bZIP, bHLH, C2C2-Dof, MYB, NAC, and WRKY), ABA signaling genes (PYR, PP2C, SnRK, and ABI5), and other genes involved in fatty acid biosynthesis (FATA, FATB, LACS, SAD, Oleosins, and so on). These results indicated that ABA treatment promoted the expression of FAD2 and other genes involved in fatty acid biosynthesis, which possibly resulted in the accumulation of linoleic acid. This study will be helpful for understanding the possible mechanisms through which ABA affects fatty acid biosynthesis and their accumulation in the mesocarp of oil palm.

An ABA Functional Analogue B2 Enhanced Salt Tolerance by Inducing the Root Elongation and Reducing Peroxidation Damage in Maize Seedlings.

Salt stress negatively affects maize growth and yield. Application of plant growth regulator is an effective way to improve crop salt tolerance, therefore reducing yield loss by salt stress. Here, we used a novel plant growth regulator B2, which is a functional analogue of ABA. With the aim to determine whether B2 alleviates salt stress on maize, we studied its function under hydroponic conditions. When the second leaf was fully developed, it was pretreated with 100 µM ABA, 0.01 µM B2, 0.1 µM B2, and 1 µM B2, independently. After 5 days treatment, NaCl was added into the nutrient solution for salt stress. Our results showed that B2 could enhance salt tolerance in maize, especially when the concentration was 1.0 µMol·L−1. Exogenous application of B2 significantly enhanced root growth, and the root/shoot ratio increased by 7.6% after 6 days treatment under salt stress. Compared with control, the ABA level also decreased by 31% after 6 days, which might have resulted in the root development. What is more, B2 maintained higher photosynthetic capacity in maize leaves under salt stress conditions and increased the activity of antioxidant enzymes and decreased the generation rate of reactive oxygen species by 16.48%. On the other hand, B2 can enhance its water absorption ability by increasing the expression of aquaporin genes ZmPIP1-1 and ZmPIP1-5. In conclusion, the novel plant growth regulator B2 can effectively improve the salt tolerance in maize.

Abscisic Acid in Coordination with Nitrogen Alleviates Salinity-Inhibited Photosynthetic Potential in Mustard by Improving Proline Accumulation and Antioxidant Activity

This investigation was done to assess the role of abscisic acid (ABA; 25 µM) and/or nitrogen (N; 10 mM) in the alleviation of salinity (NaCl; 100 mM)-induced reduction in photosynthetic activity and growth, N and sulfur (S) assimilation of mustard (Brassica juncea L.) cv. RH0-749. Salinity treatment caused oxidative stress and significantly elevated the content of both H2O2 and thiobarbituric acid reactive substances (TBARS), and impaired photosynthetic activity and growth, but increased the content of nitrogenous osmolyte proline and the activity of antioxidant enzymes involved in the metabolism of reactive oxygen species. The application of 25 µM ABA under a controlled condition negatively affected photosynthesis and growth. However, ABA, when combined with N, minimized oxidative stress and mitigated the salinity-inhibited effects by increasing the activity of antioxidant enzymes (superoxide dismutase, SOD; glutathione reductase, GR; ascorbate peroxidase, APX) and proline content. Overall, the supplementation of 10 mM N combined with 25 µM ABA provides an important strategy for enhancing the photosynthetic potential of B. juncea under saline conditions.

Identification and characterisation of monovalent cation/proton antiporters (CPAs) in Phyllostachys edulis and the functional analysis of PheNHX2 in Arabidopsis thaliana

Plant monovalent cation/proton antiporters (CPAs), types of transmembrane transporters, play important roles in resistance to salt stress. In this study, 37 CPA genes from moso bamboo (Phyllostachys edulis) were identified and characterised. The expression profiles of 10 CPA1 genes (PheNHXs) of moso bamboo were detected by qRT-PCR, which showed that they were specifically expressed in six tissues. In addition, the expression of 10 PheNHXs in leaves and roots changed significantly under 150/200 mM NaCl and 100 μM ABA treatments. In particular, the expression of PheNHX2 in leaves and roots was significantly upregulated under NaCl treatment, thus, we cloned PheNHX2 and analysed its function. Subcellular localisation analysis showed that PheNHX2 was located on the vacuolar membrane. Overexpression of PheNHX2 reduced seed germination and root growth of Arabidopsis thaliana under salt stress, as well as severely affecting cellular Na+ and K+ content, which in turn reduced the salt tolerance of transgenic Arabidopsis. Measurements of physiological indicators, including chlorophyll content, malondialdehyde content, peroxidase and catalase enzyme activities and relative electrical conductivity, all supported this conclusion. Under salt stress, PheNHX2 also inhibited the expression of some stress-related and ion transport-related genes in transgenic Arabidopsis. Overall, these results indicate that overexpression of PheNHX2 reduces the salt tolerance of transgenic Arabidopsis. This investigation establishes a foundation for subsequent functional studies of moso bamboo CPA genes, and it provides a deeper understanding of PheNHX2 regulation in relation to the salt tolerance of moso bamboo.

Characterisation, expression and functional analysis of PAL gene family in Cephalotaxus hainanensis

Phenylalanine ammonia lyase (PAL) is the first committed step in the formation of phenylpropanoids, and catalyses the deamination of L-phenylalanine (L-Phe) to yield cinnamic acid. While PALs are common in plants, PAL genes involved in alkaloid biosynthesis in Cephalotaxus hainanensis have never been described. To obtain better knowledge of PAL genes and their number and function involved in Cephalotaxus alkaloid biosynthesis four PAL genes were screened and cloned. In vitro enzymatic analysis showed that all four PAL recombinant proteins could convert L-Phe to product trans-cinnamic acid, and showed strict substrate specificity. Moreover, the expression profiles of four ChPALs were analysed using qRT-PCR, which showed that they had higher transcript levels in roots and stems, and that different ChPALs displayed different response sensitivities and change patterns in response to stimuli. Several metabolic compounds were measured in stimulated leaves using UPLC-MS, and indicating the concentration of Cephalotaxus alkaloids and cinnamic acid in leaves subjected to different conditions. These concentrations increased significantly after treatment with 100 mM NaCl, 100 mM mannitol, 100 μM SA and 10 μM ABA. The expression levels of four PAL genes showed indications of upregulation after treatment. These results supply an important foundation for further research on candidate genes involved in the biosynthesis of Cephalotaxus alkaloids.

The Technique of Water Extract Preparation from Goat Willow Leaves with Allowance for Circadian Rhythm of Their Biological Activity to Stimulate Scots Pine Seed Germination

The preparative technique for water extraction of goat willow (Salix caprea L.) growing leaves was elaborated in order to stimulate Scots pine (Pinus sylvestris L.) seed germination by the extracts (patent no. 2662999). It was shown that the effects of the extract were accounted for by its biological activity, which considerably varied depending on the time of day for extract sampling. The maximal increase in gibberellin-like activity (equivalent to 3.4–3.6 µM GA3) and marked decrease in ABA-like activity (equivalent to 0.15‒0.30 µM ABA) were recorded in the willow leaf extracts harvested in the morning (8:00 a.m.) or evening (8:00 p.m.) in South Karelia (60° N) in mid-May. Application of the extracts to pine seeds afforded the maximal effects on their germination. Thus, the second-class seeds displayed qualitative characteristics of the first-class seeds (germination exceeding 80%) after the treatment. It is concluded that the introduction of the method into practice will allow utilization of the accessible, widespread plant raw material for production of the effective natural preparation, which is capable of improving sowing qualities of Scots pine seeds.

Abiotic stress and tissue-specific reference genes for quantitative reverse transcription PCR analysis in Korean native watermelons, Citrullus lanatus ‘Black-King’ and ‘Speed-Plus-Honey’

A wide variety of research on watermelon has been conducted, and such studies have been motivated by the published genome sequence database of watermelon. Screening of proper reference genes is the primary step for normalization in gene expression analysis. Based on previous studies conducted on Arabidopsis and cucumber, we selected eight candidate reference genes of ClACT, ClEF1α, ClGAPDH, ClIDH, ClLUG, ClPTB, ClUBC2, and Cl18SrRNA, respectively, encoding β-Actin, elongation factor 1-α, glyceraldehyde-3-phosphate-dehydrogenase, NADP-isocitrate dehydrogenase, leunig, polypyrimidine tract-binding protein1, ubiquitin-conjugating enzyme E2, and 18S ribosomal RNA from watermelon (Citrullus lanatus). The expression levels of these eight genes were evaluated by RT-qPCR under plant hormone-treatment (100 μM ABA) and abiotic stresses such as drought, cold (4 °C), and high salt concentration (250 mM NaCl). The expression patterns of these eight genes were further compared across different types of watermelon tissues such as flower, leaf, tendril, stem, root, and whole seedling. Our results showed that expressions of ClACT and ClEF1α, respectively in the Korean native watermelon cultivars Citrullus lanatus ‘Black-King’ and ‘Speed-Plus-Honey’ were least affected by the environmental stresses regardless of tissue types. Here, we suggest two ideal reference genes for watermelon RT-qPCR-based gene expression study.

EFFECT OF VARIOUS PRECULTURE AND OSMOTIC DEHYDRATION CONDITIONS ON CRYOPRESERVATION EFFICIENCY AND MORPHOGENETIC RESPONSE OF CHRYSANTHEMUM SHOOT TIPS

The aim of this study was to evaluate the influence of sucrose concentration in the preculture medium, as well as the duration of osmotic dehydration on the efficiency of chrysanthemum ‘Richmond’, ‘Lady Orange’ and ‘Lady Salmon’ cryopreservation by encapsulation-dehydration technique. For all cultivars tested, the best regrowth of cryopreserved shot tips was recorded with 0.25 M sucrose concentration and 10 µM ABA during a two-week preculture, followed by a 4- or 7-day osmotic dehydration. The survival rate ranged from 56.8–58.0% (the Lady group) to 63.6% (‘Richmond’). However, the ability to grow was smaller and reached 18.2–50.7%. It was found that higher sucrose concentration during the preculture slowed the growth of chrysanthemum shoot tips and led to an increased formation of multiple shoots (by activating axillary buds) or deformed adventitious shoots (incapable of further growth). The frequency of tissue hyperhydricity also increased, while the rhizogenesis efficiency decreased when higher sucrose concentration in the preculture medium was applied. The influence of osmotic dehydration duration on the explants morphogenetic response was cultivar-dependent

Morphogenetic response of shoot tips to cryopreservation by encapsulation-dehydration in a solid mutant and periclinal chimeras of Chrysanthemum\xa0\xd7\xa0grandiflorum /Ramat./Kitam.

Cryopreservation is widely applied to many economically important species excluding chimera plants which are problematic for long-term conservation. Their storage problems can be circumvented only by cryopreserving meristems. This study looked at the morphogenetic response of shoot tips of periclinal chimera chrysanthemum ‘Lady Orange’ and ‘Lady Salmon’, as well as the solid mutant ‘Richmond’, that were cryopreserved by encapsulation-dehydration technique. By applying 10 µM ABA in the preculture medium followed by 4-day-long dehydration treatment, the explant survival reached up to 67%. Besides the stimulation of typical single shoot recovery, cryopreservation led to direct or indirect multiple shoot formation, shoot malformation, as well as inhibited their spontaneous rooting. Microscopic analysis revealed three types of structural damages of shoot tips which can correspond with their morphogenetic response in recovery culture. No influence of cryostorage on the acclimatisation efficiency of the recovered chrysanthemums was observed.

Exploring the role of Inositol 1,3,4-trisphosphate 5/6 kinase-2 (GmITPK2) as a dehydration and salinity stress regulator in Glycine max (L.) Merr. through heterologous expression in E. coli

Phytic acid (PA) is implicative in a spectrum of biochemical and physiological processes involved in plant stress response. Inositol 1,3,4, Tris phosphate 5/6 kinase (ITPK), a polyphosphate kinase that converts Inositol 1,3,4 trisphosphate to Inositol 1,3,4,5/6 tetra phosphate, averting the inositol phosphate pool towards PA biosynthesis, is a key regulator that exists in four different isoforms in soybean. In the present study, in-silico analysis of the promoter region of ITPKs was done and among the four isoforms, promoter region of GmITPK2 showed the presence of two MYB binding elements for drought inducibility and one for ABA response. Expression profiling through qRT-PCR under drought and salinity stress showed higher expression of GmITPK2 isoform compared to the other members of the family. The study revealed GmITPK2 as an early dehydration responsive gene which is also induced by dehydration and exogenous treatment with ABA. To evaluate the osmo-protective role of GmITPK2, attempts were made to assess the bacterial growth on Luria Broth media containing 200 mM NaCl, 16% PEG and 100 μM ABA, individually. The transformed E. coli BL21 (DE3) cells harbouring the GmITPK2 gene depicted better growth on the media compared to the bacterial cells containing the vector alone. Similarly, the growth of the transformed cells in the liquid media containing 200 mM NaCl, 16% PEG and 100 μM ABA showed higher absorbance at 600 nm compared to control, at different time intervals. The GmITPK2 recombinant E. coli cells showing tolerance to drought and salinity thus demonstrated the functional redundancy of the gene across taxa. The purity and specificity of the recombinant protein was assessed and confirmed through PAGE showing a band of ∼35 kDa on western blotting using Anti- Penta His- HRP conjugate antibody. To the best of our knowledge, the present study is the first report exemplifying the role of GmITPK2 isoform in drought and salinity tolerance in soybean.

ABA Is Involved in Regulation of Cold Stress Response in Bermudagrass.

As a representative warm-season grass, Bermudagrass [Cynodon dactylon (L). Pers.] is widely used in turf systems. However, low temperature remarkably limits its growth and distribution. ABA is a crucial phytohormone that has been reported to regulate much important physiological and biochemical processes in plants under abiotic stress. Therefore, the objective of this study was to figure out the effects of ABA on the cold-sensitive (S) and cold-resistant (R) Bermudagrass genotypes response to cold stress. In this study, the plants were treated with 100 μM ABA solution and exposed to 4°C temperature. After 7 days of cold treatment, the electrolyte leakage (EL), malonaldehyde (MDA) and H2O2 content were significantly increased in both genotypes compared with control condition, and these values were higher in R genotype than those of S genotype, respectively. By contrast, exogenous ABA application decreased the electrolyte leakage (EL), MDA and H2O2 content in both genotypes compared with those plants without ABA treatment under cold treatment condition. In addition, exogenous ABA application increased the levels of chlorophyll a fluorescence transient curve for both genotypes, and it was higher in R genotype than that of S genotype. Analysis of photosynthetic fluorescence parameters revealed that ABA treatment improved the performance of photosystem II under cold condition, particularly for the R genotype. Moreover, cold stress significantly increased δ13C values for both genotypes, while it was alleviated by exogenous ABA. Additionally, exogenous ABA application altered the expression of ABA- or cold related genes, including ABF1, CBF1, and LEA. In summary, exogenous ABA application enhanced cold resistance of both genotypes by maintaining cell membrane stability, improving the process of photosystem II, increasing carbon isotopic fractionation under cold stress, and more prominently in R genotype compared with S genotype.