Showing papers on "Zeatin published in 2020"

Exogenous Brassinolide Enhances the Growth and Cold Resistance of Maize (Zea mays L.) Seedlings under Chilling Stress

Abstract: This paper aims to elucidate the effects of exogenous brassinolide (BL) on maize germination and seedling growth under chilling stress. The cold-resistant maize hybrid Tiannong 9 and the cold-sensitive hybrid Tianhe 1 were soaked at the germination stage (6 °C) and leaves were sprayed at seedling stage (4 °C), with BL at concentrations of 0, 0.01, 0.1, and 1 mg/L. The germination rate of the maize seeds and the changes in seedling biomass, antioxidant, photosynthetic, and plant endogenous hormone systems and chloroplast ultrastructures were determined. The results showed that the optimum concentration of BL to alleviate chilling stress in maize seedlings was 0.1 mg/L. This rate effectively increased the germination rate and plant biomass of maize and significantly increased the superoxide dismutase (SOD) peroxidase (POD) and catalase (CAT) activities, the net photosynthetic rate (Pn), stomatal conductance (gs) and transpiration rate (Tr), and seedling auxin (IAA), gibberellin (GA3) and trans zeatin nucleoside (t-ZR) contents under chilling stress. In addition, BL significantly reduced the malondialdehyde (MDA) content, abscisic acid (ABA) content, and intercellular carbon dioxide concentration (Ci). In the comparison of mesophyll cells, the chloroplast membrane of the treatment group was tightly attached to the stroma, and some of the plasma membranes were dissolved, but the overall structure of the chloroplast was relatively complete, and the osmiophilic granules were relatively few. The exogenous application of BL can effectively alleviate the damage caused by a low temperature in maize, maintain the normal characteristics of seedlings in chilling environments, and ensure the development and growth of plant tissue in the later stage.

trans-Zeatin-N-glucosides have biological activity in Arabidopsis thaliana.

Abstract: Cytokinin is an indispensable phytohormone responsible for physiological processes ranging from root development to leaf senescence. The term “cytokinin” refers to several dozen adenine-derived compounds occurring naturally in plants. Cytokinins (CKs) can be divided into various classes and forms; base forms are generally considered to be active while highly abundant cytokinin-N-glucosides (CKNGs), composed of a CK base irreversibly conjugated to a glucose molecule, are considered inactive. However, results from early CK studies suggest CKNGs do not always lack activity despite the perpetuation over several decades in the literature that they are inactive. Here we show that exogenous application of trans-Zeatin-N-glucosides (tZNGs, a specific class of CKNGs) to Arabidopsis results in CK response comparable to the application of an active CK base. These results are most apparent in senescence assays where both a CK base (tZ) and tZNGs (tZ7G, tZ9G) delay senescence in cotyledons. Further experiments involving root growth and shoot regeneration revealed tZNGs do not always have the same effects as tZ, and have largely distinct effects on the transcriptome and proteome. These data are in contrast to previous reports of CKNGs being inactive and raise questions about the function of these compounds as well as their mechanism of action.

Nitrogen Deficiency-Induced Decrease in Cytokinins Content Promotes Rice Seminal Root Growth by Promoting Root Meristem Cell Proliferation and Cell Elongation.

Abstract: Rice (Oryza sativa L.) seedlings grown under nitrogen (N) deficiency conditions show a foraging response characterized by increased root length. However, the mechanism underlying this developmental plasticity is still poorly understood. In this study, the mechanism by which N deficiency influences rice seminal root growth was investigated. The results demonstrated that compared with the control (1 mM N) treatment, N deficiency treatments strongly promoted seminal root growth. However, the N deficiency-induced growth was negated by the application of zeatin, which is a type of cytokinin (CK). Moreover, the promotion of rice seminal root growth was correlated with a decrease in CK content, which was due to the N deficiency-mediated inhibition of CK biosynthesis through the down-regulation of CK biosynthesis genes and an enhancement of CK degradation through the up-regulation of CK degradation genes. In addition, the N deficiency-induced decrease in CK content not only enhanced the root meristem cell proliferation rate by increasing the meristem cell number via the down-regulation of OsIAA3 and up-regulation of root-expressed OsPLTs, but also promoted root cell elongation by up-regulating cell elongation-related genes, including root-specific OsXTHs and OsEXPs. Taken together, our data suggest that an N deficiency-induced decrease in CK content promotes the seminal root growth of rice seedlings by promoting root meristem cell proliferation and cell elongation.

Endogenous Hormones Inhibit Differentiation of Young Ears in Maize (Zea mays L.) Under Heat Stress.

Abstract: Global warming frequently leads to extreme temperatures, which pose a serious threat to the growth, development, and yield formation of crops such as maize. This study aimed to deeply explore the molecular mechanisms of young ear development under heat stress. We selected the heat-tolerant maize variety Zhengdan 958 (T) and heat-sensitive maize variety Xianyu 335 (S), and subjected them to heat stress in the V9 (9th leaf), V12 (12th leaf), and VT (tasseling) growth stages. We combined analysis of the maize phenotype with omics technology and physiological indicators to compare the differences in young ear morphology, total number of florets, floret fertilization rate, grain abortion rate, number of grains, and main metabolic pathways between plants subjected to heat stress and those left to develop normally. The results showed that after heat stress, the length and diameter of young ears, total number of florets, floret fertilization rate, and number of grains all decreased significantly, whereas the length of the undeveloped part at the top of the ear and grain abortion rate increased significantly. In addition, the differentially expressed genes (DEGs) in young ears were significantly enriched in the hormone signaling pathways. The endogenous hormone content in young ears exhibited different changes: zeatin (ZT) and zeatin riboside (ZR) decreased significantly, but gibberellin acid3 (GA3), gibberellin acid4 (GA4), and abscisic acid (ABA) increased significantly, in ears subjected to heat stress. In the heat-tolerant maize variety, the salicylic acid (SA), and jasmonic acid (JA) content in the vegetative growth stage also increased in ears subjected to heat stress, whereas the opposite effect was observed for the heat-sensitive variety. The changes in endogenous hormone content of young ears that were subjected to heat stress significantly affected ear development, resulting in a reduction in the number of differentiated florets, fertilized florets and grains, which ultimately reduced the maize yield.

Naturally Occurring and Artificial N9-Cytokinin Conjugates: From Synthesis to Biological Activity and Back

Abstract: Cytokinins and their sugar or non-sugar conjugates are very active growth-promoting factors in plants, although they occur at very low concentrations. These compounds have been identified in numerous plant species. This review predominantly focuses on 9-substituted adenine-based cytokinin conjugates, both artificial and endogenous, sugar and non-sugar, and their roles in plants. Acquired information about their biological activities, interconversions, and metabolism improves understanding of their mechanisms of action and functions in planta. Although a number of 9-substituted cytokinins occur endogenously, many have also been prepared in laboratories to facilitate the clarification of their physiological roles and the determination of their biological properties. Here, we chart advances in knowledge of 9-substituted cytokinin conjugates from their discovery to current understanding and reciprocal interactions between biological properties and associated structural motifs. Current organic chemistry enables preparation of derivatives with better biological properties, such as improved anti-senescence, strong cell division stimulation, shoot forming, or more persistent stress tolerance compared to endogenous or canonical cytokinins. Many artificial cytokinin conjugates stimulate higher mass production than naturally occurring cytokinins, improve rooting, or simply have high stability or bioavailability. Thus, knowledge of the biosynthesis, metabolism, and activity of 9-substituted cytokinins in various plant species extends the scope for exploiting both natural and artificially prepared cytokinins in plant biotechnology, tissue culture, and agriculture.

Differential expression of microRNAs in tomato leaves treated with different light qualities

Abstract: Light is the main source of energy and, as such, is one of the most important environmental factors for plant growth, morphogenesis, and other physiological responses. MicroRNAs (miRNAs) are endogenous non-coding RNAs that contain 21–24 nucleotides (nt) and play important roles in plant growth and development as well as stress responses. However, the role of miRNAs in the light response is less studied. We used tomato seedlings that were cultured in red light then transferred to blue light for 2 min to identify miRNAs related to light response by high-throughput sequencing. A total of 108 known miRNAs and 141 predicted novel miRNAs were identified in leaf samples from tomato leaves treated with the different light qualities. Among them, 15 known and 5 predicted novel miRNAs were differentially expressed after blue light treatment compared with the control (red light treatment). KEGG enrichment analysis showed that significantly enriched pathways included zeatin biosynthesis (ko00908), homologous recombination (ko03440), and plant hormone signal transduction (ko04075). Zeatin biosynthesis and plant hormone signal transduction are related to plant hormones, indicating that plant hormones play important roles in the light response. Our results provide a theoretical basis for further understanding the role of miRNAs in the light response of plants.

Temporal and spatial expression analysis of shoot-regeneration regulatory genes during the adventitious shoot formation in hypocotyl and cotyledon explants of tomato (CV. Micro-Tom)

Abstract: Enhancing the competence for plant regeneration in tissue culture studies is an important issue not only for efficient genetic transformation of commercial crops but also for the reproducibility of scientific reports. In this study, we investigated optimization of several tissue culture conditions including plant growth regulators, types and ages of explants, culture densities, and plant position in order to improve the competence of adventitious shoot formation of the tomato (Solanum lycopersicum cv. Micro-Tom). In addition, we examined the differential expression of D-type cyclin (CYCD3-1) and several shoot regeneration regulatory genes from hypocotyl and cotyledon explants of tomato during shoot organogenesis. A treatment of 1 mg L-1 Zeatin and 0.1 mg L-1 Indole-3-acetic acid (IAA) in Murashige and Skoog (MS) medium containing 3% sucrose was optimal for adventitious shoot formation from hypocotyl and cotyledon explants. The younger explants exhibited more shoot formation regardless of explant types. Additionally, those closest to the shoot apical meristem produced more shoots compared to the other regions in the hypocotyl and the cotyledon explants. Gene expression of CYCD3-1, SHOOT MERISTEMLESS (STM), and cytokinin dependent WUSCHEL (WUS) was significantly higher in younger explants than in older ones. Furthermore, an increase in CYCD3-1, STM, and WUS expression was evident at the distal part of hypocotyls and the proximal part of cotyledons compared to other regions. These differential gene expression profiles exhibited good agreement with the results of shoot formation obtained from diverse explants of tomato. These results suggest that temporal and spatial gene expression of shoot regeneration regulatory genes plays an important role in enhancing the competence and the reproducibility of adventitious shoot formation from tomato explants.

Endogenous Phytohormones of Fern Polystichum aculeatum (L.) Roth Gametophytes at Different Stages of Morphogenesis in vitro Culture

Abstract: The content of indole-3-acetic (IAA), gibberellic (GA3), abscisic acids (ABA), and cytokinins—zeatin (Z), zeatin glucoside (ZG), zeatin riboside (ZP), isopentenyladenin (iР), and isopentenyladenosine (iРА)—was determined using the method of high-performance liquid chromatography in fern Polystichum aculeatum gametophytes at different stages of morphogenesis in vitro culture. It was shown that, at the stage of spatulate prothallium development, gametophytes contained significant amounts of GA3 and IAA, while the content of ABA and cytokinins was low. The inactive ZG and active iP were dominant. At the stage of cordiform thallus formation, the level of GA3 increased by 1.6, IAA by 2.4, and ABA by 1.7 times. The content of inactive iPA increased twice, significantly increased the level of active ZR and iP. At the development stage of sporophyte on the surface of gametophyte thallus, the levels of IAA, ABA, and GA3 decreased. The accumulation of inactive ZG and decrease in the content of active zeatin and zeatin riboside were documented. At all stages of gametophytes’ morphogenesis, GA3 was dominant, and its highest content was detected during the development of archegonia and antheridia. The pattern of gibberellic acid and zeatin riboside accumulation has shown the key role of these hormones in the regulation of growth processes and the development of reproductive structures of P. aculeatum gametophytes.

Effects of Gibberellin (GA4+7) in Grain Filling, Hormonal Behavior, and Antioxidants in High-Density Maize (Zea mays L.).

Abstract: Dense plant cultivation is an efficient approach to improve maize production by maximizing the utilization of energy and nutrients. However, dense plant populations may aggravate the abortion rate of young grains, resulting in fewer kernels per ear. The rate and duration of grain-filling play decisive roles in maize grain yield. Therefore, to increase plant density, enhancing the grain-filling rate, extending the growth period of individual maize plants and regulating crop senescence would be the first priority. In this study, we examined the regulatory effects of GA4+7 under two application methods: shanks and silks were moistened by cotton full with GA4+7 solution at concentrations of 0, 10, 60, and 120 mg L-1. The results showed that GA4+7 improved the grain-filling rate by increasing the content of auxin, gibberellin, zeatin, and abscisic acid in grains compared to control plants. In addition, the auxin, gibberellin, and zeatin contents in the grains were positively and significantly correlated with the maximum grain weight and the maximum and mean grain-filling rates. Moreover, GA4+7 increased the activities of superoxide dismutases, catalases, and peroxidases and reduced the malondialdehyde content in leaves compared with untreated plants. At the concentration of 60 mg L-1, GA4+7 showed the greatest effect on shank and silk applications (Sh-60 and Si-60) followed by 10 mg L-1 (Sh-10) for shank treatment and 120 mg L-1 (Si-120) for silk treatment. Our results suggest that a concentration of 60 mg L-1 GA4+7 for shank and silk application may be efficiently used for changing the level of hormones in grains and antioxidant enzymes in ear leaves, which may be useful for enhancing grain-filling rate and delaying leaf senescence, resulting in an increase in maize grain yield.

Advancements in Low-Chill Blueberry Vaccinium corymbosum L. Tissue Culture Practices

Abstract: The demand for blueberry Vaccinium corymbosum L. (and hybrids) plants has significantly increased in the last 30 years due to its market expansion. In vitro propagation of sterile plants are required for commercial purposes but also for research applications such as plant transformation. Thus far, tissue culture characteristics of the tropical-adapted blueberry have been scarcely studied. In this study we present the following findings: (i) zeatin, a hormone used to promote plant growth, should be used in the 1–2 mg/L range to promote plant architecture optimal for transformation experiments; (ii) red-blue LED lights induce more production of meristems and biomass than white LED or fluorescent lights; (iii) levels as high as 1000 mg/L of decontamination agents (the antibiotics timentin and cefotaxime) can be used to eliminate Agrobacterium overgrowth without inhibiting plant growth during plant transformation experiments; (iv) kanamycin, paromomycin, and geneticin, which are widely used antibiotics to select transgene-carrying transformants, cannot be efficiently used in this system; (v) glufosinate, a widely used herbicide, shows potential to be used as an effective selectable marker for transformed plants.

Rapid generation of transgenic and gene-edited Solanum nigrum plants using Agrobacterium-mediated transformation

Abstract: In this study, genetic engineering methods, ranging from gene transformation to gene editing, were efficiently conducted on cotyledon explants in Solanum nigrum. Organogenic calli were observed on the explants following 10 days of growth on medium supplemented with 2 mg/L zeatin and 0.2 mg/L indole acetic acid (IAA), which we suggest to be an efficient shoot initiation medium (SM1). In vitro shoot production was enhanced in explants grown on media supplemented with 1 mg/L zeatin and 0.1 mg/L IAA, which we used as a proper shoot differentiation medium (SM2) in S. nigrum shoot regeneration. Direct infection of explants with Agrobacterium harboring CRISPR/Cas9 constructs can simplify the method of Agrobacterium-mediated T-DNA transformation by skipping the preincubation step. The method was applied to deliver transgenes for genome editing, such as CRISPR/Cas9 DNA. SnLazy1 locus, considered to be orthologs of tomato Lazy1, was edited using the CRISPR/Cas9 system in S. nigrum. Two independent deletion snlazy1-cr alleles were successively inherited and showed stem growth in a relatively downward direction. Our method offers rapid and simple procedure for gene transformation and genome editing that could be applicable for the enhancement of beneficial traits for precision plant breeding and engineering quantitative trait loci in S. nigrum.

Seed germination, micropropagation from adult and juvenile origin explants and address of hyperhydricity of the Cretan endemic herb Calamintha cretica

Abstract: The optimum range of temperature for germination (96-100%) of Calamintha cretica, an herb with potential pharmaceutical and horticultural uses, was 15 to 20 °C, with 10 and 30 °C cardinal temperatures. Storage up to one year did not affect germination. The effect of zeatin (ZEA), 6-benzyladenine (BA), kinetin, and 6-γ-γ-(dimethylallylamino)-purine added in MS medium at concentrations from 0.0 to 8.0 mg L-1 was tested for shoot proliferation of both adult- and seedling-origin nodal explants at first- and sub-culture. Both explant types responded similarly during in vitro culture. At cytokinin concentrations up to 1 mg L-1 explant response was high (over 85%) but shoot number per explant was low (1.2-2.2). Increasing cytokinin from 2.0 to 8.0 mg L-1 resulted to an analogous decrease of explant response and shoot length, and an increase of shoot number, particularly when ZEA or BA was used (5.0-6.6 shoots per explant, 0.5-1.0 cm long) with simultaneous though increase of hyperhydricity (up to 50%). The addition of 0.1 mg L-1 naphthaleneacetic acid into the 8.0 mg L-1 BA medium almost eliminated hyperhydricity and increased explant response, while the increase of agar concentration from 8.0 to 12.0 g L-1 eliminated hyperhidricity and induced the highest shoot proliferation (93-95% explant response, 11.2-12.3 shoots per explant, 0.8-1.0 cm long). Microshoots and microshoot clusters rooted (88-96%) on half-strength MS medium either hormone free or supplemented with 1 to 4 mg L-1 indole-3-butyric acid. Plantlets survived at 80% to 100% after ex vitro acclimatization in peat: perlite 1:1 (v/v).

Comparative regeneration in six bread wheat (Triticum aestivum L.) varieties from immature and mature scutella for developing efficient and genotype-independent protocol prerequisite for genetic improvement of wheat

Abstract: Hexaploid bread wheat, being highly recalcitrant, has been difficult to regenerate in vitro which limits the use of transgenesis and genome editing for its agronomic improvement and functional genomics. We have studied the comparative in vitro regeneration in six most important wheat varieties, namely, HD-2967, PBW-343, WH-1105, PBW-550, DBW-17, and WL-711, using immature vs mature scutellum. The regeneration response of varieties PBW-550, DBW-17, and HD-2967 has not yet been reported. The mature and immature scutella cultured on MS medium containing 2,4-dichlorophenoxyacetic acid (2,4-D) (2 mg l−1) or dicamba (6 mgl−1), respectively, and maltose (30 gl−1) under dark conditions developed calli which subsequently on transfer to modified MS medium with double concentration of NH4NO3, zeatin (5.0 mgl−1), dicamba (0.1 mgl−1), AgNO3 (2.0 mgl−1), CuSO4 (2.5 mgl−1), and sucrose (30 gl−1) under light conditions regenerated into shoots. Immature scutellum showed 100% callus induction irrespective of varieties and regenerated a maximum of 15 shoots per callus in 90% cultures, whereas mature scutellum showed a maximum 80% callus induction, producing maximum 5 shoots only in 67% cultures. Thus, immature scutellum has been more responsive comparative to mature scutellum for inducing callus and shoot regeneration. Among all the six varieties, HD-2967 was the most regenerative, which can be explored further for improving wheat via genetic transformation studies.

Effect of microalgae Messastrum gracile and Chlorella vulgaris on the in vitro propagation of orchid Cattleya labiata

Abstract: The aim of this study was to evaluate the effect of two green microalgae (Messastrum gracile and Chlorella vulgaris) in comparison with plant growth regulators (6-benzylaminopurine, BAP; thidiazuron, TDZ; zeatin, ZEA) on the in vitro propagation of Cattleya labiata, an endangered orchid, using the thin cell layer (TCL) technique from protocorms. TCL sections were cultivated in MS/2 medium containing M. gracile extract (EM) and biomass (BM); C. vulgaris extract (EC) and biomass (BC); and BAP, TDZ, and ZEA in different concentrations. Subsequently, the explants were grown in MS/2 medium, with 2 g L−1 of activated charcoal, to induce elongation and roots formation. For acclimatization, plants were transplanted in trays using Sphagnum sp. as substrate. TCL explants showed a higher formation of protocorm-like bodies (PLBs) than entire protocorms. Explants cultivated in media supplemented with BM showed a high rate of PLB regeneration (59%) and high mean number of PLBs (4 per explant) and 85% survival after acclimatization of the plants. Supplementation with BAP stimulated similar morphogenic responses to those observed with BM and superior results obtained with ZEA and TDZ. Mass propagation of C. labiata plants was successfully achieved using TCL, and it is recommended to supplement the MS/2 medium with 4 g L−1 of BM or 4 mg L−1 of BAP. Microalgae extracts and biomasses are effective alternatives for in vitro propagation of C. labiata that can replace plant growth regulators, as they favored the formation of PLBs and plants.

Morphogenesis, pigment content, phytohormones and productivity of eggplants under the action of gibberellin and tebuconazole

Abstract: One of the main tasks of modern plant physiology is regulation of growth and development of cultivated plants in order to optimize the productive process. The attention of the scientific community is focused on the use of natural activators and growth inhibitors. We investigated the effect of foliar treatment with 0.005% solution of gibberellic acid (GA3) and 0.025% solution of the antigibberellic preparation tebuconazole (EW-250) on morphogenesis, leaf mesostructure, the content of photosynthetic pigments, the balance of endogenous phytohormones and productivity of Capsicum annuum L., Antey variety. The vegetation experiment was carried out in the conditions of soil-sand culture in vessels with a volume of 10 L. Treatment of plants was carried out in the budding phase. Morphometric parameters were determined every 10 days. The mesostructure of the middle tier leaves was studied in the fruit formation phase, and the chlorophyll content was determined in the raw material by spectrophotometric method. Analytical determination of endogenous phytohormones – indolyl-3-acetic (IAA), gibberellic (GA3) and abscisic (ABA) acids and cytokinins – zeatin (Z), zeatin-O-glucoside (ZG), zeatinribozide (ZR), isopentenyladenine (iP) and isopentenyladenosine (iPA) were performed by high performance liquid chromatography – mass spectrometry (HPLC-MS). With GA3 treatment, plant height increased considerably, while with EW-250, it decreased. Both regulators led to an increase in the number of leaves on the plant, the leaf raw biomass, stems and roots and the dry matter of the whole plant, the area of a single leaf blade and the total area of leaves on the plant. Under the action of EW-250, the chlorophyll content in the leaves surged, while under the action of GA3 it tended to decrease or did not change at all. Both regulators thickened the chlorenchyma and boosted the volume of the columnar parenchyma cells. GA3 treatment induced a rise in the thickness of the upper and lower epidermis, and EW-250 led, on the contrary, to a decrease. It is shown that after treatment with exogenous GA3, the content of endogenous IAA and ABA decreased and GA3 in plant stems increased. Instead, EW-250 caused a decrease in the levels of GA3, IAA and ABA in the stems. Exogenous GA3 enhanced the accumulation of endogenous GA3 and IAA and inhibited ABA in the leaves. Under the action of the retardant, the level of ABA in the leaves did not change, while GA3 and IAA decreased. Treatment of plants with the studied growth regulators caused a decrease in the pool of cytokinins (CK) in stems. EW-250 showed a significant rise in the hormone content in the leaves. After spraying with GA3 solution, the level of ZG, Z and ZR grew. Under the action of the retardant, the increase in the CK pool occurred exclusively due to the iP. Growth regulators optimized the productivity of sweet pepper plants: under the action of GA3 there was an increase in the number of fruits per plant, and after the use of EW-250 there was a rise in the average weight of one fruit. The obtained results showed that anatomical-morphological and structural-functional rearrangements of sweet pepper plants under the action of exogenous gibberellic acid and EW-250 took place against the background of changes in the balance and distribution of endogenous hormones. Increased photosynthetic activity, stimulation of growth processes of some plant organs and inhibition of others enlarged biological productivity of the culture.

High frequency direct shoot regeneration from Kazakh commercial potato cultivars.

Abstract: Potato (Solanum tuberosum L.) is the third most economically important crop in the world and has a high nutritional value. In this study, the in vitro culture response of four widely grown in Kazakhstan potato cultivars, Astanalyk, Monument Kunaev, Tokhtar, and Aksor, was investigated using stem and leaf explants. Published protocols were evaluated and optimized to develop a more efficient protocol for the regeneration of plants from local potato cultivars in tissue culture, which is a prerequisite to facilitate potato genome modification. The explants were cultured on solid Murashige and Skoog medium supplemented with different concentrations and combinations of zeatin, 6-benzylaminopurine (BAP), gibberellic acid (GA3), 1-naphthaleneacetic acid (NAA) and indole-3-acetic acid (IAA). The maximum regeneration was induced from the stem internodal explants. A significant effect of the explant source on direct regeneration was confirmed with statistical analysis. The number of shoots obtained from the internode was 10.0 from cv. Aksor followed by cvs. Tokhtar and Astanalyk. The medium DRM-VIII with 1 mg/l zeatin, 0.1 mg/l IAA and 7.0 mg/l GA3 was considered the best for direct shoot regeneration and multiple shoot formation from all cultivars. To conclude, we outline a protocol for direct plant regeneration from four potato cultivars. Our findings suggest commercial cultivars Astanalyk and Aksor are good candidates for developing the genome-edited plants through direct shoot regeneration.

Transcriptome analyses provide insights into the homeostatic regulation of axillary buds in upland cotton (G. hirsutum L.).

Abstract: The axillary bud is an important index of cotton plant-type traits, and the molecular mechanism of axillary bud development in upland cotton has not yet been reported. We obtained a mutant (designated mZ571) with a high-budding phenotype in axillary bud development from the low-budding phenotype variety G. hirsutum Z571 (CCRI 9A02), which provided ideal materials for the study of complex regulatory networks of axillary bud development. In this study, RNA sequencing was carried out to detect gene expression levels during three stages of axillary buds in Z571 (LB, low budding) and mZ571 mutant (HB, high budding). A total of 7162 DEGs were identified in the three groups (HB-E vs. LB-E, HB-G1 vs. LB-G1, HB-G2 vs. LB-G2), including 4014 downregulated and 3184 upregulated DEGs. Additionally, 221 DEGs were commonly identified in all three groups, accounting for approximately 3.09% of the total DEGs. These DEGs were identified, annotated and classified. A significant number of DEGs were related to hormone metabolism, hormone signal transduction, and starch and sucrose metabolism. In addition, 45, 22 and 9 DEGs involved in hormone metabolic pathways and 67, 22 and 19 DEGs involved in hormone signal transduction pathwayspathway were identified in HB-E vs. LB-E, HB-G1 vs. LB-G1, and HB-G2 vs. LB-G2, respectively, suggesting that endogenous hormones are the primary factors influencing cotton axillary bud growth. Hormone and soluble sugar content measurements revealed that mZ571 exhibited higher concentrations of zeatin, gibberellins and soluble sugar in all three stages, which confirmed that these hormone metabolism-, hormone signal transduction- and starch metabolism-related genes showed interaction effects contributing to the divergence of axillary bud growth between mZ571 and Z571. Our results confirmed the importance of endogenous hormones and sugars in the development of axillary buds, and we found that mZ571 plants, with a high-budding phenotype of axillary buds, exhibited higher endogenous hormone and sugar concentrations. Overall, we present a model for the emergence and development of cotton axillary buds that provides insights into the complexity and dynamic nature of the regulatory network during axillary bud emergence and development.

Comparative transcriptome analyses provide novel insights into etiolated shoot development of walnut (Juglans regia L.)

Abstract: In general, genes promoting IAA, CTK GA and ethylene biosynthesis were upregulated, while genes participating in ABA, chlorophyll and starch biosynthesis pathways performed opposite tendency during etiolation. Etiolation as a method for rejuvenation plays an important role in the vegetative propagation of woody plants. However, the molecular mechanism of etiolated shoot development remains unclear. In this study, we investigated changes at different etiolation stages of Juglans regia. The histology and transcriptome of J. regia were analysed using etiolated stems, which were treated in darkness for 30, 60, 90 days. The results showed that the ratios of pith (Pi) diameter/stem diameter (D), cortex (Co) width/D, and phloem (Ph) width/D increased, while the ratio of xylem (Xy) width/D decreased after etiolation, and the difference in these ratios between etiolated stems and the control was more significant at 60 days than 90 days. Differentially expressed genes (DEGs) were significantly enriched in pathways such as plant hormone biosynthesis and signal transduction, chlorophyll biosynthesis and degradation, and starch and sucrose metabolism. The difference in the contents of indole-3-acetic acid (IAA), abscisic acid (ABA), sugar and chlorophyll between etiolated stems and the control increased with increasing treatment duration; in contrast, the concentrations of gibberellin (GA), zeatin (ZT), and starch, as well as the difference between the etiolated stems and control were lowest at 60 days among the three stages. On the whole, the positive effect of etiolation on the rejuvenation of walnut stems changed as the treatment period increased. The present investigation lays a foundation for future studies on the effect of etiolation on rejuvenation and for promoting the efficiency of vegetative propagation.

Changes in essential oil composition, phenylalanine ammonia lyase gene expression and rosmarinic acid content during shoot organogenesis in cytokinin-treated Satureja spicigera (C. Koch) boiss. shoots

Abstract: This study investigated the effects of different concentrations of zeatin and thidiazuron on the accumulation of phenolics and activity of phenylalanine ammonia lyase gene in the micropropagated Satureja spicigera shoots. S. spicigera clones were generated from cultured nodal segments on Murashige and Skoog including vitamins medium supported with zeatin and thidiazuron concentrations. Shoots were subcultured three times in the same medium and harvested at the end of the third month. The expression levels of phenylalanine ammonia lyase gene were conducted using semi-quantitative reverse transcription polymerase chain reaction technique. Rosmarinic acid contents were determined using reverse phase-high performance liquid chromatography with a diode array detector. Genetic stability of regenerants was performed using random amplified polymorphic DNA markers. Essential oils from shoots were identified by gas chromatography–mass spectrophotometry analysis. It has been detected that 0.1 mg/L thidiazuron is more effective on the node formation and shoot elongation. Contrarily, 4.0 mg/L thidiazuron caused a significant increase in shoot multiplication. The highest biomass accumulation was calculated from medium including 1.0 mg/L zeatin. According to random amplified polymorphic DNA analysis, all banding profiles from micropropagated plants were monomorphic. Gas chromatography–mass spectrophotometry analysis showed that rising concentrations of zeatin and thidiazuron increased the thymol and decreased the carvacrol content. It was also found that rosmarinic acid production increased significantly at 1.0 mg/L zeatin application. However, thidiazuron applications had no positive effect on rosmarinic acid accumulation. Comparing phenylalanine ammonia lyase gene expression and rosmarinic acid accumulation in in vitro shoots, it was found that they were correlated.

Efficient direct shoot organogenesis and genetic stability in micropropagated sacha inchi (Plukenetia volubilis L.).

Abstract: It is necessary to improve biotech platforms based on in vitro cell tissue culture to support sacha inchi (Plukenetia volubilis L.) research programs and draw on the nutritional value of the high polyunsaturated fatty acid content of its oilseed. Here, we developed a rapid and efficient method for induction and direct in vitro shoot development for this species. Shoots were generated from hypocotyl explants. The highest organogenic response was obtained in woody plant medium supplemented with 1 mg/L thidiazuron and 0.5 mg/L zeatin supplemented with L-glutamine, adenine hemisulfate, and L-arginine. Shoots obtained using this medium were transferred and subcultivated with different concentrations of indole-3-butyric acid and 1-naphthylacetic acid for rooting. For the first time, a histological analysis was performed supporting direct organogenic development in this species. The plantlets obtained were transferred ex vitro with a survival percentage of 80%. The genetic stability of the plants recovered was confirmed by randomly amplified polymorphic DNA analysis. All results indicate that it would be possible to stimulate direct shoot formation from hypocotyls to support the sustainable use of this species.

Effects of Cytokinin on in vitro Propagation of Bauhinia variegata L.

Abstract: Mature seeds of Bauhinia variegata L were cultured on half strength Murashige and Skoog medium. For experimentation, nodal cuttings were used as explants from in vitro growing plants. Cytokinin, N-benzyl-9-(2-tetrahydropyranyl) (BPA), kinetin(6-furfurylaminopurine), zeatin, 6-(4-hydroxy-3-methyl-trans -2-butenyl amino purine), 2- isopentenyl amino purine (2-ip), and benzylaminopurine (BAP) were tested for best propagation. Well grown plants were achieved in medium supplemented with 5 µM BPA and 0.5 µM BAP. The propagated plants were acclimatized very well after transferred to the field.

Multiple shoot induction and plant regeneration from axillary buds of Magnolia ‘Vulcan’

Abstract: An efficient protocol for multiple shoot induction and plant regeneration from axillary bud culture of Magnolia ‘Vulcan’ was developed in the present study. Primary shoots were obtained from axillary bud explants cultured on Murashige and Skoog (MS) medium containing 1.0 mg/L 6-benzylaminopurine (BA). To induce multiple shoots effectively, primary shoot tips were cultured on MS medium supplemented with different concentrations of BA and zeatin at 0, 0.2, 0.5, and 1.0 mg/L. Of these treatments, the MS medium with 0.5 mg/L BA resulted in the highest number of shoots per explant with an average value of 5.9, and it produced the greatest shoot height at 4.8 cm after 12 weeks of culturing. In the rooting of in vitro produced shoots, the greatest percentage of explants forming roots (91.3%), number of roots per explant (9.7), and root length (2.8 cm) were obtained in half-strength MS medium supplemented with 6.0 mg/L indole-3-butyric acid (IBA). Regenerated plantlets were successfully acclimatized and hardened off inside the culture room with 87.5% survival rate. Plants were transferred to a greenhouse with a 97.2% survival rate. The highly efficient shoot multiplication and plant regeneration system reported herein can be used for large-scale clonal propagation of valuable Magnolia species or cultivars.