Micropropagation of Daylily (<i>Hemerocallis fulva</i>) from Crown-Tip Explants and Assessment of Somaclonal Variation of <i>in Vitro</i>-Propagated Plants Using SCoT Markers

Determination of the somaclonal variation of in vitro-propagated plants is crucial to determine the appropriatemicropropagation protocol and growth regulators for commercial scale multiplication. In this research, nine multiplication media (MM) augmented with different concentrations of 6-benzyl adenine (BA), Kinetin (Kin), andThidiazuron (TDZ), Three rooting media (RM) supplemented with three levels of α-naphthalene acetic acid(NAA) and three types of soil mixtures (v/v); Coco peat/Vermiculite/Sand (CVS), Peat moss/Perlite/Sand(PPS) and Peat moss/Perlite (PP) were used in the micropropagation protocol of daylily plants. MM2 showedthe maximum shoot length and the number of leaves, while MM9 showed the maximum number of shoots.The RM1 showed the maximum root length and the number of roots. During acclimatization, CVS, PPS, andPP soil mixture showed similar performance except the CVS mixture showed lower performance regarding plantheight and diameter. The genetic fidelity of micropropagated plants was evaluated using Start Codon Targeted(SCoT) Markers. Six SCoT primers amplified 51 scorable bands with an approximate range from 146 bp to1598 bp size. Thirty one out of 51 loci were presented in the mother plants. 40 loci were polymorphic, 11 weremonomorphic and 7 were unique. The amplification patterns of the micropropagated plants demonstrated genetic integrity to the mother plant ranging from 84.32 to 47.06 and somaclonal variations ranging from 52.94 with5 mg/l BA pathway to 15.68 with 1mg/l TDZ pathway, thus demonstrating that the homogeneity and the variation of the micropropagated plants affected by the type and the quantity of the plant growth regulator used duringmultiplication subcultures. This research can be successfully used for other ornamental and medicinal plants’ bulkmultiplication, germplasm conservation, and future genetic improvement.     

Morphometric and Biochemical Changes in <i>Agave americana</i> L. Plantlets Induced By Ethyl Methanesulfonate

A. americana L. is a crop with very little genetic variability. In order to evaluate the effect of ethyl methanesulfonate (EMS) to induce variability in in vitro plantlets of A. americana, different explants (meristems, leaves and roots) were evaluated for the production of callus. MS medium supplemented with ANA (2.68 μM) and BAP (2.68 μM) was used. Callus obtained from apical meristem were treated with 15 mM EMS for two hours after which shoot formation was induced using 2,4-D (0.11 μM) and BAP (44 μM). The EMS induced variations in the morphometric and morphological parameters of the plantlets obtained, with 60% of the plantlets presenting differences such as dwarfism and different leaf forms, without the presence of spines, as well as an increase in fructan content of 30% with respect to the control plantlets. PAL was increased and this activity is related with higher anthocyanins concentration in A. americana L. plantlets.     

Antifungal Potential of <i>Beauveria bassiana</i> on <i>Solanum lycopersicum</i> L. Infected with <i>Fusarium oxysporum</i> f. sp. <i>lycopersici</i>

The objective of this work was to evaluate the effect of Beauveria bassiana (Bb 1205) on controlling Fusarium oxysporum f. sp. lycopersici (Fol 17108) in tomato plants in greenhouse conditions. Inoculation of Bb 1205 was the most promising among the agronomic variables and expression of the activity of the enzymes β-1,3-glucanases and chitinases. Inoculation of Bb 1205 occurred at a concentration of 1 × 10⁸ conidia·mL⁻¹, which was administered onto the leaves, directly into the soil and via injection. Infection with Fol 17108 occurred with 1 × 10⁶ spores·mL⁻¹, which were added directly to the soil. Spectrophotometry was used for measuring agronomic parameters, namely activity of chitinases and β-1,3-glucanases in foliage and roots. When Bb 1205 was added to the soil, the chlorophyll index and aerial part length showed significant differences. In addition, it was determined that root length, fresh weight of foliage, flower, and fruit count increased 82 days after inoculation (dai). Chitinase activity induced by Bb 1205 in leaves and roots of tomato plants infected with Fol 17108 was observed when injected into the stem at 32 dai (41.8 and 11.6-fold, respectively). Inoculation on the foliage showed a 10-fold increase of β-1,3-glucanases in the roots after 82 dpi. As for leaves, a 3.8-fold increase was found when the stem was inoculated. In the different in vivo applications, Bb 1205 activated its defenses by expressing the chitinase enzymes and β-1,3-glucanase, thus reducing the damage caused by Fol 17108, demonstrating increase plant growth thereafter.     

Effect of Water Stress Induced by Polyethylene Glycol on Growth,Proline Accumulation in <i>Agave americana</i> L.

The effect of water deficit was determined on both in vitro and soil seedling as well as in cells in suspension of Agave americana L. In order to do the establishment of cells, the formation of callus was induced; for it two auxins were evaluated: 2,4-dichlorophenoxyacetic acid (2,4-D) and 4-mino-3,5,6-trichloropicolinic acid (picloram) at three concentrations (0.25, 0.5 and 0.75 mg L⁻¹) in three explants (leaf, root and meristems) cultured in MS semisolid medium. The callogenesis response was related to the type and section of the explant, as well as the regulator used, and a cell suspension was established using 0.5 mg L⁻¹ naphthaleneacetic acid (NAA) + 0.5 mg L⁻¹ Benzylaminopurine (BAP). Seedlings were exposed to polyethyleneglycol (15% and 30% w/v) with a water potential of −0.87 and −2.67 MPa, respectively, under soil conditions. Water stress was applied through restricted irrigation. Fresh weight, root system growth, and chlorophyll concentration were some of the parameters that were affected by the effect of water deficit on A. americana L. Chlorophyll concentration values were significantly decreased by 15 at 30% PEG (19.6 SPAD units) compared to the control treatment. In in vitro plants, the highest concentration of proline was found in the roots, being the treatment with 30% polyethylene glycol where the highest concentration of this osmoregulator was obtained (62.5 mg g⁻¹ DW). Under restricted irrigation conditions, an increase in proline concentration was observed both in the aerial part (2.2 µg 100 g⁻¹ DW) and in the root system (1.8 µg 100 g⁻¹ DW). However, the concentrations found were approximately ten times greater, less than those found under in vitro conditions. Therefore, the accumulation of proline can be considered an indicator of stress in Agave Americana L. growth in vitro.     

Pre-Breeding Genetic Diversity Assessment of Tomato (<i>Solanum lycopersicum</i> L.) Cultivars Based on Molecular,Morphological and Physicochemical Parameters

Appropriate knowledge of the parental cultivars is a pre-requisite for a successful breeding program. This study characterized fruit yield, quality attributes, and molecular variations of ten tomato cultivars during three consecutive generations under greenhouse conditions. Peto 86, Castle Rock, and Red Star cultivars showed the highest fruit yield (kg/plant), total phenolic compounds (TPC), and sap acidity. Principal component analysis categorized the evaluated fruit yield into three groups based on their quality attributes. A robust positive correlation appeared among traits inside each group. A positive correlation was likewise noticed between the first and the second groups. However, a negative correlation was detected between the first, the second and the third group. Molecular profiling, using seven inter-simple sequence repeat (ISSR) primers, produced 60 loci, including 49 polymorphic loci. The molecular analysis also pinpointed the highest genetic similarity (0.92) between P73 and Moneymaker, while the lowest genetic similarity (0.46) was observed between Castle Rock and Moneymaker. The cultivars P73 and Moneymaker showed the lowest genetic distance (2.24), while the highest genetic distance (5.92) was observed between Super Marmand and Peto86, on the one hand, and between Castle Rock and Moneymaker, on the other hand. The chemical analysis of fruit sap indicated the highest levels of TPC, total flavonoids, anthocyanin, ascorbic acid and total soluble solids in Peto 86 and Castle Rock cultivars. Phylogeny analysis of tomato cultivars based on morphological and molecular attributes indicated four distinct clades. Peto 86, Castle Rock, and Red star cultivars can be recommended for the tomato hybridization breeding programs in the future, with other tomato cultivars as potentially high-yielding parents.     

Molecular Interaction Study to Explore the <i>Nigella sativa</i> Bioactive Components as an Inhibitor of Peptide Deformylase to Inhibit the <i>Xanthomonas oryzae</i> pv. oryzae via Applying Computational Approach

Bacterial leaf blight (BLB) caused by Xanthomonas oryzae pv. oryzae (Xoo) is one of the most damaging diseases to rice across the world. Various chemicals have been employed so far for the management of bacterial leaf blight. On the other hand, these compounds are damaging to the ecosystem and have an impact on non-target species such as humans and animals. As a result, there is a need to create a new natural inhibitor for BLB management. Deformylase (PDF) enzyme is present in all eubacteria and its necessity in bacterial protein synthesis reveals it as an attractive target for drug development. In this study, the active components of Nigella sativa have been selected based on their previously reported antimicrobial activity and screened on the active site of bacterial PDF by the in silico art of techniques. Among these compounds, dithymoquinone and p-cymene strongly bind with the PDF enzyme with binding energy values of 7.77 kcal/mol and 7.26 kcal/mol, respectively, which is comparatively higher than the control compound (−6.73 kcal/mol). Hence, the “dithymoquinone-PDF” and “p-cymene-PDF” complexes were selected for further study, and their stability was assessed by molecular dynamic (MD) simulation. In MD simulation, both selected compounds exhibited steady-state interaction with PDF for 20 ns. It has been hypothesized that p-cymene and dithymoquinone inhibit peptide deformylase and could be used as antibacterials or pesticides against Xoo against the BLB disease.     

<i>In Vitro</i>-Propagation of <i>Agave tequilana</i> Weber cv. azul in a Temporary Immersion System

In Mexico, there is a need to produce large quantities of plantlets for the establishment and replanting of blue (cv. azul) agave production areas. Most of these plots are within the origin denomination area (DOT, Spanish acronym) of the distilled product of this plant, known as tequila. The objective of this study was to develop an in vitro-propagation protocol for Agave tequilana Weber cv. azul using segmented stems in both: solid and liquid media. A disinfection and in vitro technique were developed to obtain shoots, through plantlets collected in commercial plots, which attained 100% surface-disinfection and budding rate. At the multiplication stage, the effects of 6-Benzylaminopurine (BA) (0.0, 4.4 and 13.2 μM) and kinetin (0.0, 9.4, 18.8 and 37.6 μM) were evaluated on lateral-shoot production of segmented sagittal stems. These were cultivated on Murashige & Skoog (MS) medium, with the addition of 3.0% sucrose and 8 g L⁻¹ agar. It was observed that BA and kinetin increased the number of shoots per explant, obtaining up to 18 and 26, respectively. Furthermore, it was found that just the sagittal segmentation of explants increased axillary budding. On the other hand, segmented-stem bases were grown in MS liquid medium with 3.0% sucrose, inside a RITA® system, programmed by a 5 min immersion step with a frequency of every 4 h. The effect of Indole−3-Acetic acid (IAA) (0.57, 2.9, 5.7 μM) was evaluated, while maintaining a concentration of BA (13.2 μM). It was observed that the greatest concentration of IAA led to the formation of more than 20 buds per explant. These results offer a new methodology to increase the efficiency of A. tequilana Weber cv. azul-in vitro multiplication by sagittal segmentation of stems and the addition of BA and/or IAA.     

<i>In Vitro</i> and <i>in Silico</i> Insights on the Biological Activities,Phenolic Compounds Composition of <i>Hypericum perforatum</i> L. Hairy Root Cultures

Three Hypericum perforatum hairy root lines (HR B, HR F and HR H) along with non-transformed roots were analyzed for phenolic compounds composition and in vitro enzyme inhibitory properties. In silico molecular modeling was performed to predict the interactions of the most representative phenolic compounds in HR clones with enzymes related to depression, neurodegeneration and diabetes. Chromatographic analyses revealed that HR clones represent an efficient source of quinic acid and hydroxybenzoic acids, epicatechin and procyanidin derivatives, quercetin and kaempferol glycosides, as well numerous xanthones. In vitro antidepressant activity of HR extracts through monoamine oxidase A (MAO-A) inhibition was attributed to the production of oxygenated and prenylated xanthones. The neuroprotective potential of HR extracts was related to the accumulation of quercetin 6-C-glucoside, epicatechin, procyanidins and γ-mangostin isomers as potential inhibitors of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). Vanillic acid and prenylated xanthones in HR clones as promising inhibitors of tyrosinase additionally contributed to the neuroprotective activity. Five preeminent xanthones in HR (γ-mangostin, mangiferin, garcinone C, garcinone E and 1,3,7-trihydroxy-6-metoxy-8-prenyl xanthone) along with the flavonol quercetin 6-C-glucoside effectively inhibited α-amylase and α-glucosidase indicating the antidiabetic properties of HR extracts. Transgenic roots of H. perforatum can be exploited for the preparation of novel phytoproducts with multi-biological activities.     

Genetic Improvement of <i>Betula platyphylla</i> Suk. in China: A Review

Birch (Betula platyphylla Suk.), distributed in Eurasia, North America, and Australia, is a kind of cold-resistant, fast-growing, and vital pulpwood tree species. It is also one of the most important ecological restoration tree species with high values of economic benefits in Northeast China. To improve the genetic gain and expand the economic benefit of B. platyphylla, many genetic improvements have been carried out. In China, B. platyphylla is widely distributed and varied, and there are many varieties with excellent genetic characteristics. In this paper, the genetic improvement of B. platyphylla was reviewed, and the previous research results were discussed from two aspects: conventional breeding and molecular breeding. Some problems and corresponding solutions in the genetic improvement were put forward to provide ideas for B. platyphylla breeding in the future.     

Genetic Variation in Growth and Cone Traits of <i>Pinus Koraiensis</i> Half-Sib Families in Northeast China

Genetic parameters were evaluated for growth and cone characteristics (tree height, diameter at breast height, volume, cone number, thousand seeds weight and single cone seeds weight) on 86 half-sib families of Pinus koraiensis aged 31 years. Analyses of variance revealed significant differences (p < 0.001) in all growth and cone traits among families while no significant differences were detected among blocks and the interaction between blocks and families. The average family values for growth traits were 17.22 m, 8.67 cm and 0.43 m3 for tree height, diameter at breast height and volume, respectively. The average cone number, thousand seeds weight and single cone seeds weight were 17.57, 748.91 g and 77.25 g, respectively. Genotypic additive variance and phenotypic variances ranged from 0.00009 to 3.820 and from 0.0005 to 23.066, while genotypic and phenotypic coefficients of variation ranged from 2.693% to 37.196% and 4.963% to 60.595%, respectively. Heritability at the individual and family level ranged from 0.152 to 0.215 and 0.611 to 0.862, respectively. Growth traits were significantly positively correlated with each other, but cone traits showed a weak correlation with growth traits. Based on 10% selection rate, nine families each were selected as elite materials in terms of high performance in volume and cone numbers, with 22.16% and 43.82% genetic gain in volume and cone number, respectively. These results provide beneficial information to select excellent families and establish orchards of P. koraiensis from improved seeds.     

G × E Analysis of Growth Traits of <i>Betula platyphylla</i> Clones at Three Separated Sites in Northeastern China

To select elite materials, the growth traits of 32 Betula platyphylla clones at three separated northern sites in Northeast China were investigated and analyzed. The results showed that there were significant differences among all variation sources in the different investigated traits (P < 0.01). Except for the carbon contents, all the coefficients of phenotypic variation of the other investigated traits were higher than 10%. The repeatability of different traits ranged from 0.760 to 0.998. Correlation analysis showed that tree height were significantly correlated with diameter at breast height, but neither was significantly correlated with leaf traits nor element contents. Additive main effects and multiplicative interaction analysis showed that genotype, environment and genotype × environment interactions were significantly different in diameter at breast height, which indicated that environment had a significant effect on genotype. Comprehensive assessment results showed that three clones with high and stable diameters at breast height were selected, and the genetic gains of diameter at breast height on sites Maoershan, Qingan, and Yongji were 21.24%, 20.58%, and 38.65%, respectively. The results could provide a theoretical basis for elite clone selection in B. platyphylla and other broad leaved species.     

Cloning and Characterization of <i>EuGID1</i> in <i>Eucommia ulmoides</i> Oliver

Gibberellic acid controlled the key developmental processes of the life cycle of landing plants, and regulated the growth and development of plants. In this study, a novel gibberellin receptor gene EuGID1 was obtained from Eucommia ulmoides Oliver. The cDNA of EuGID1 was 1556 bp, and the open reading frame was 1029 bp, which encoded 343 amino acids. EuGID1 had the homology sequence with the hormone-sensitive lipase family. Amino acid sequence alignment confirmed EuGID1 protein had the highest homology with the GID1 protein of Manihot esculenta. EuGID1 was located in the nucleus and cell membrane and had expression in four plant organs. Overexpression of EuGID1 in transgenic Arabidopsis plants promoted plant elongation and increased siliques yield.     

Factors influencing in the response of <i>Schizolobium parahybum</i> (Vell) Blake to <i>Ceratocystis paradoxa</i> and <i>C. moniliformis</i>

In the Ecuadorian coast one of the most destructive diseases of the pachaco is vascular wilt or stem rot caused by Ceratocystis complex, so the aim of this study was to determine the factors that affect the efficiency of the reaction of bark pachaco to this disease. This research was conducted under laboratory conditions, using trees pachaco S38, S41, S98, AE-1, AE-2 and AE-3, and pathogenic species Ceratocystis paradoxa and C. moniliformis. The method utilized was tissue stem bark,with bark sections with 4.5 cm², and a suspension of 3x10⁴ units infection and remained in a humid chamber for 96 hours at 25 ± 5 °C. Were determined grades of resistance/ susceptibility using a scale from 0 to 4, depending on the amount of mycelia and peritecio in each plant sample. Three factors were used: four colonies obtained by several transfers from each fungal specie, four ages of colonies of each fungal specie and four volumes of inoculum applied (units of infection), using for each experiment separately Completely Randomized Design with 4 replications factorial arrangement. For comparison between treatment means was used Tukey test at 5% probability of error. For future trials using this technique, you could use 30-day colonies for C. paradoxa and 40 days for C. moniliformis, and an application volume of 100 μL/cm², it would improve the level of response for the formation of perithecium and mycelia in samples cortex.     

Differential Responses of <i>NHX1</i> and <i>SOS1</i> Gene Expressions to Salinity in two <i>Miscanthus sinensis</i> Anderss. Accessions with Different Salt Tolerance

The lignocellulosic crop Miscanthus spp. has been identified as a good candidate for biomass production. The responses of Miscanthus sinensis Anderss. to salinity were studied to satisfy the needs for high yields in marginal areas and to avoid competition with food production. The results indicated that the relative advantages of the tolerant accession over the sensitive one under saline conditions were associated with restricted Na⁺ accumulation in shoots. Seedlings of two accessions (salt-tolerant ‘JM0119’ and salt-sensitive ‘JM0099’) were subjected to 0 (control), 100, 200, and 300 mM NaCl stress to better understand the salt-induced biochemical responses of genes involved in Na⁺ accumulation in M. sinensis. The adaptation responses of genes encoding for Na⁺ /H⁺ antiporters, NHX1 and SOS1 to NaCl stress were examined in JM0119 and JM0099.The cDNA sequences of genes examined were highly conserved among the relatives of M. sinensis based on the sequencing on approximate 600 bp-long cDNA fragments obtained from degenerate PCR. These salt-induced variations of gene expression investigated by quantitative real-time PCR provided evidences for insights of the molecular mechanisms of salt tolerance in M. sinensis. The expression of NHX1 was up-regulated by salt stress in JM0119 shoot and root tissues. However, it was hardly affected in JM0099 shoot tissue except for a significant increase at the 100 mM salt treatment, and it was salt-suppressed in the JM0099 root tissue. In the root tissue, the expression of SOS1 was induced by the high salt treatment in JM0119 but repressed by all salt treatments in JM0099. Thus, the remarkably higher expression of NHX1 and SOS1 were associated with the resistance to Na⁺ toxicity by regulation of the Na⁺ influx, efflux, and sequestration under different salt conditions.     

Selection and Verification of Reference Genes for qRT-PCR Analysis in <i>Iris domestica</i> under Drought

Iris domestica is a plant of the Iridaceae family and is drought-tolerant, but its drought-resistance mechanism is not yet clear. Analysing the gene expression changes of I. domestica by qRT-PCR is an important mean to understand its drought resistance characteristics. Nevertheless, a lack of reference genes greatly hinders investigation and research on the adaptation of I. domestica to drought at the molecular and genetic levels. In this study, we assessed the expression stability of 11 candidate gene in I. domestica under drought stress conditions and different tissues using geNorm, NormFinder, BestKeeper and RefFinder tools. The results showed that EF1β was the most stable reference genes under drought stress and in different tissues. To validate further the stability of the identified reference genes, the expression patterns of VP gene in I. domestica was analysed. These results will be conducive to more accurate quantification of gene expression levels in I. domestica.