Axillary Shoot Research Articles

Natural Variation in the Control of Flowering and Shoot Architecture in Diploid Fragaria Species.

In perennial fruit and berry crops of the Rosaceae family, flower initiation occurs in late summer or autumn after downregulation of a strong repressor TERMINAL FLOWER1 (TFL1), and flowering and fruiting takes place the following growing season. Rosaceous fruit trees typically form two types of axillary shoots, short flower-bearing shoots called spurs and long shoots that are, respectively, analogous to branch crowns and stolons in strawberry. However, regulation of flowering and shoot architecture differs between species, and environmental and endogenous controlling mechanisms have just started to emerge. In woodland strawberry (Fragaria vesca L.), long days maintain vegetative meristems and promote stolon formation by activating TFL1 and GIBBERELLIN 20-OXIDASE4 (GA20ox4), respectively, while silencing of these factors by short days and cool temperatures induces flowering and branch crown formation. We characterized flowering responses of 14 accessions of seven diploid Fragaria species native to diverse habitats in the northern hemisphere and selected two species with contrasting environmental responses, Fragaria bucharica Losinsk. and Fragaria nilgerrensis Schlecht. ex J. Gay for detailed studies together with Fragaria vesca. Similar to F. vesca, short days at 18°C promoted flowering in F. bucharica, and the species was induced to flower regardless of photoperiod at 11°C after silencing of TFL1. F. nilgerrensis maintained higher TFL1 expression level and likely required cooler temperatures or longer exposure to inductive treatments to flower. We also found that high expression of GA20ox4 was associated with stolon formation in all three species, and its downregulation by short days and cool temperature coincided with branch crown formation in F. vesca and F. nilgerrensis, although the latter did not flower. F. bucharica, in contrast, rarely formed branch crowns, regardless of flowering or GA20ox4 expression level. Our findings highlighted diploid Fragaria species as rich sources of genetic variation controlling flowering and plant architecture, with potential applications in breeding of Rosaceous crops.

Establishment of In Vitro Regeneration Protocol for Sabah’s Jewel Orchid, Macodes limii J.J. Wood & A.L. Lamb

Habitat disturbance and excessive collection of wild orchids from their natural habitat have threatened many orchids species at risk of extinction. In this study, the in vitro regeneration protocol for Macodes limii, a jewel orchid endemic to Sabah was established. The effects of explant source and plant growth regulators (PGRs) including naphthaleneacetic acid, picloram, 2,4-dichlorophenoxyacetic acid, 6-benzylaminopurine, kinetin, and thidiazuron on the in vitro regeneration capacity of M. limii plantlets were examined. Both factors showed a significant interaction in promoting axillary shoot formation. Nodal explants from the third and fourth positions cultured with 1.0 mg/L TDZ, induced 95% of shoot regeneration, with an average of three shoots/explant (1.6–1.8 cm of shoot length) after 90 days of culture. The well-developed plantlets went through an acclimatization phase for 60 days with a 60% of survival rate. An inter simple sequence repeat (ISSR) marker analysis confirmed the genetic stability of the in vitro regenerated plants to the mother plant. The successfully acclimatized plantlets were finally transferred to Poring Orchid Conservation Centre for reintroduction. The established protocol provides the means for large-scale production of this endemic jewel orchid, as well as a basis for further research aimed at the conservation and genetic improvement of this plant.

In-Vitro Growth Enhancement of Cedrela odorata L. using Benzyl Amino Purine and Α-Naphthalene Acetic Acid

Studies were conducted in order to develop protocol for mass-production of Cedrela odorata seedlings. Clean-plantlets were initiated on four media at full and half strengths. Shoot-regeneration consisted two media types (Murashige & Skoog (MS) and Woody Plant Medium (WPM)) and four Benzyl Amino Purine (BAP) concentrations: 0.0, 1.0, 2.0 and 3.0 mg/l. Root-induction involved three MS media strengths (25, 50 and 100 % basal salts) and two Naphthalene Acetic Acid (NAA) levels (0.0 and 0.5 mg/l) all in factorial combinations and laid out in completely randomised design (CRD). The results at initiation showed that MS at full strength enhanced in vitro seeds growth better in terms of root lengths (6.50 cm) and number of leaves (7.6) while WPM gave better shoot length (6.24 cm) at 3 Week after inoculation (WAI). Shoots were best regenerated on MS basal medium supplemented with 1.0 mg/l BAP for shoot length at 4 WAI while the effects of BAP levels were comparable but higher than control for number of leaves and axillary shoots. Highest number of roots (3.92) induced and root length (2.93 cm) obtained were from 100% MS medium at 8 WAI. Hence, MS medium (100% basal salt) with 1.0 mg/l BAP with or without 0.5 mg/l NAA are considered suitable protocol for in vitro propagation of C. odorata.

Efficient micropropagation of Thunbergia coccinea Wall. and genetic homogeneity assessment through RAPD and ISSR markers

Thunbergia coccinea Wall. ex D. Don being a rare, ornamental and medicinal plant of India, is needed to propagate for conserving the germplasm and analyzing its phytochemical compounds in the future. A reliable protocol for direct in vitro propagation using nodal shoot meristem of T. coccinea as explant was standardized. The highest number of shoots per explant (22.17 ± 0.54) with maximum shoot length (2.36 ± 0.28) in cm was obtained in Murashige and Skoog (MS) medium supplemented with 9.70 µM of 6-furfurylaminopurine (Kinetin) and 0.053 µM of α-naphthaleneacetic acid (NAA) combination, among all the different plant growth regulators (PGR’s) and concentrations tested. The aforesaid PGR’s combination was optimum for axillary shoot bud induction and multiplication in T. coccinea. The best rooting was observed on the half-strength MS medium fortified with 2.68 µM NAA with the highest number of roots per shoot (3.75 ± 0.12) and maximum length (5.22 ± 0.32) in cm. All the in vitro raised plantlets were acclimatized in sterile sand and soil mixture (1:1) with a survival rate of 70% on earthen pots under greenhouse conditions. PCR-based RAPD (Random Amplified Polymorphic DNA) and ISSR (Inter-Simple Sequence Repeat) molecular markers were employed to determine the genetic homogeneity amongst the plantlets. Twelve (12) RAPD and nine (9) ISSR primers developed a total of 104 and 91 scorable bands, respectively. The band profiles of micropropagated plantlets were monomorphic to the mother, donor in vivo plant, and similarity values varied from 0.9542–1.000. The dendrogram generated through UPGMA (unweighted pair group method with arithmetic mean) showed 99% similarities amongst all tested plants confirming the genetic uniformity of in vitro raised plants.

In vitro propagation and ex vitro rooting of Euonymus verrucosus Scop. (Celastraceae) – a rare species of Kazakhstan flora on the southern border of its areal

ABSTRACT Euonymus verrucosus Scop. (Celastraceae) is a boreal European-Mediterranean-Asia Minor forest relict species and is listed in the Red Data Books of Kazakhstan and some regions of the Russian Federation. Thus, in the present study, in vitro regeneration of E. verrucosus utilizing young shoot explants was attempted for the first time. Initially, the young shoots (0.8–1.0 cm length) were placed in woody plant medium (WPM) and Murashige and Skoog’s (MS) medium fortified with different concentrations of cytokinins, such as 6-benzylaminopurine (BAP), kinetin (Kn) and thidiazuron (TDZ), to induce axillary buds. The optimum induction was 75.0 ± 2.9% and the apical growth was 3.74 ± 0.06 cm for young shoots after 4 weeks in WPM medium supplemented with BAP (1.0 mg L−1). Simultaneously, during the formation of axillary shoots, callus induction was observed at the nodal base. Subculturing this callus on fresh medium every 4 weeks generated total six adventitious shoots with 0.70 ± 0.02 cm length after 12 weeks in the presence of BAP (1.0 mg L−1). When the adventitious shoots were isolated and transferred again to WPM medium containing 2.0 mg L−1 Kn, 2.95 ± 0.16 shoots were formed with 51.2% regeneration frequency within 4 weeks. These microshoots were then rooted ex vitro and an optimum 73.3% rooting was achieved within 4 weeks in a substrate highbog peat with a 0.5 cm layer of vermiculite. This micropropagation protocol will be helpful for the conservation of the genetic resources and biodiversity for E. verrucosus.

Validation of meta-Topolin in organogenesis, improved morpho-physio-chemical responses, and clonal fidelity analysis in Dioscorea pentaphylla L. – an underutilized yam species

The present investigation describes the efficacy of aromatic and synthetic cytokinins, which influence the direct organogenesis, biochemical responses, and ex vitro and in vivo performance of the in vitro propagated plantlets of Dioscorea pentaphylla L., an underrated famine tuber crop. The axillary shoot bud induction was achieved on the Murashige and Skoog's (MS) medium supplemented with 1.0 mg L−1 meta Topolin (mT) (96.9%) and 1.0 mg L−1 Thidiazuron (TDZ) (82.4%). The sprouted shoots were proliferated using 0.5 mg L−1 mT combined with 0.1 mg L−1 indole-3-acetic acid (IAA) with the maximum average number of 43.0 ± 1.29 shoots (7.2 ± 0.55 cm length). The shoots proliferated on mT and IAA combination demonstrated the presence of higher concentrations of carbohydrates and photosynthetic pigment contents in the leaves as compared with TDZ and IAA derived leaves. The healthy micro-shoots were rooted on a half-strength MS medium containing 1.0 mg L−1 indole-3-butyric acid (IBA) with the development of 12.6 ± 0.38 roots (5.9 ± 0.22 cm length). The plantlets were acclimatized in a greenhouse and showed 100% survival under in vivo conditions. The clonal fidelity of the regenerants was evaluated for the first time using Start codon targeted (SCoT) markers. Thirteen primers amplified 62 well-resolved bands and ranged from 1-8 bands. There were no somaclonal variations detected among the regenerants of D. pentaphylla with the mother plant.

Effect of antioxidants and growth regulators on shoot organogenesis in the apical meristem culture of <i>Fragaria × ananassa</i> (Duchesne ex Weston) Duchesne ex Rozier

The initiation of strawberries into in vitro culture is known to be complicated by the inhibition of organogenesis by phenolic oxidation products. An important role in this process is given to the selection of growth regulators that increase meristematic cell activity and shoot proliferation at the stage of organogenesis induction. The present study aims to obtain a viable apical meristem culture of garden strawberry and to study the effect of different antioxidants (reduced glutathione (RG); a new preparation, i.e., a mechanical composite (MC) on the basis of biogenic silicon and green tea catechins and plant growth regulators (6-benzylaminopurine; thidiazuron) on the initiation of axillary shoot formation in strawberry meristem culture. Terminal buds containing an apical meristem and two leaf primordia isolated from the stolons of two garden strawberry cultivars (Sunny Meadow and Festival Chamomile) were used as primary explants for the initiation of strawberries into in vitro culture. It was found for the first time that the MC exhibits higher antioxidant activity as compared to reduced glutathione, reduces darkening of initial explants, as well as enhancing regeneration up to 13.0% at p ≤ 0.05. Furthermore, the best effect on the formation of microshoots per explant is observed toward the end of material introduction into in vitro culture when combining the MC with growth regulators in the culture medium. Here, the effect of strawberry cultivar on explant regeneration and the number of microshoots per explant are insignificant. It is concluded that the procedure for using the MC as an effective antioxidant during material initiation into the culture can be applied to the large-scale in vitro propagation of garden strawberries. Moreover, the technology for obtaining the MC from plant waste is environmentally friendly, which is a significant advantage for its use in in vitro technologies.

Spontaneous Regeneration of Plantlets Derived from Hairy Root Cultures of Lopezia racemosa and the Cytotoxic Activity of Their Organic Extracts.

A histological analysis was performed with the aim of elucidating the spontaneous regeneration process of the hairy root lines LRT 2.3 and LRT 6.4, derived from Lopezia racemosa leaf explants and genetically transformed with the Agrobacterium rhizogenes strain ATCC15834/pTDT. The analysis showed both lines regenerate via indirect somatic embryogenesis; LRT 6.4 also regenerated by direct organogenesis. The morphogenic characteristics of the regenerated plantlets from both lines showed the typical characteristics, described previously, including a higher number of axillary shoot formation, short internodes, and plagiotropic roots compared with wild-type seedlings. The regeneration process occurred without the addition of plant growth regulators and was linked to the sucrose concentration in the culture medium. Reducing the sucrose concentration from 3% to 2%, 1%, and 0.5% increased the regeneration rate in LRT 6.4; the effect was less pronounced in LRT 2.3. The cytotoxic activity of different organic extracts obtained from roots and shoots were evaluated in the cancer cell lines HeLa (cervical carcinoma), HCT-15 (colon adenocarcinoma), and OVCAR (ovary carcinoma). The hexane and dichloromethane extracts from roots of both lines showed cytotoxic activity against the HeLa cell line. Only the dichloromethane extract from the roots of PLRT 2.3 showed cytotoxic activity against the OVCAR cell line. None of the methanol extracts showed cytotoxic activity, nor the shoot extracts from any solvent.

In-Vitro Regeneration of Interspecific F1 Hybrid (Eucalyptus citriodora and Eucalyptus torelliana) of Eucalyptus

Tissue culture technique was standardized for in vitro shoot multiplication, using nodal segments of 25- 30 years old trees i.e., promising interspecific F1 hybrid of Eucalyptus (Eucalyptus citriodora and Eucalyptus torelliana). Due to the phenolic exudation explants did not survive and eventually died without regenerating buds. The explants collected from July to September were the best for in vitro studies for icropropagation. The axillary buds were surface sterilized with 0.1% Mercuric chloride solution for 10-15 minutes, followed by 0.1% Bavistin treatment for 1 minute and subsequently washed 3-4 times with sterilized distilled water. These surface sterilized axillary buds were cultured on MS medium supplemented with cytokinin and auxin. MS medium supplemented with 1.5mg/l BAP + 0.1mg/l NAA proved to be the best hormonal combination for induction of axillary bud which resulted in the development of 1-3 axillary shoots. The proliferated shoots were cultured on MS medium with different concentration of BAP (0.1 – 3.0 mg/l) alone or in combination with NAA (0.1-1.5mg/l) and supplemented with sucrose at 3%. The aim of the paper was to obtain maximum rate of in vitro regeneration of FRI-15 by using micropropagation technique.

In vitro shoot regeneration of Milicia excelsa (Welw.) C.C. Berg

The need for mass propagation and genetic breeding of tree species with economic importance, such as Milicia excelsa, requires an in vitro regeneration protocol. Different concentrations of Gibberellic acid (GA3) (0, 1, 2, 3 and 4 mg/L) and 6-benzylaminopurine (BAP) (0.0, 0.5, 1.0, 1.5 and 2 mg/L) were added to Murashige and Skoog medium for seed germination and shoot regeneration of the species. The treatments were laid out in completely randomised design. Each set of GA3 and BAP treatments were composed by 20 and 6 replicates, respectively. Observations made on cultured seeds include germination percentage at 4 weeks interval while shoot length, number of shoots, number of roots and leaves collected on regenerated shoots at 2 weeks interval were subjected to analysis of variance and means separated with Fisher’s protected Least Significant Difference (L.S.D) at p ≤ 0.05. Results showed that using GA3 positively influenced the seed germination as early germination occurred in the media with GA3, in comparison to the treatment without plant growth regulator (control). Nonetheless, 2.0 mg/L GA3 was observed to be the optimum, having the highest germination percentage of 30% at 4 Weeks after Inoculation (WAI). The results of the BAP effects on the shoot regeneration showed that 0.5 mg/L was the optimum for best shoot growth, having the average number of leaves (6.5), axillary shoots (1.8) and highest shoot length (3.75 cm) in comparison to the control treatment. In conclusion, these results provide insight on in vitro propagation of Milicia excelsa and the techniques employed could be adopted for its shoot multiplication. Index terms: In vitro seed germination; micropropagation; multiple shoots; moraceae.

Some clonal micropropagation features ofLonicera caeruleaL. cultivarsin vitro

Lonicera caeruleaL. is one of the early temperate berry crops. Honeysuckle is a deciduous shrub up to 2.5 m high with edible blue fruits rich in vitamins and polyphenols. The work is devoted to the improvement ofin vitropropagation method of someLonicera caeruleacultivars. The differences in regeneration and formation of axillary shoots were revealed depending on the mineral medium composition. The optimal mineral composition of the nutrient medium for clonal micropropagation of the studied genotypes was determined. Cultivation on of Quoirin and Lepoivre medium contributed to the maximum reproduction coefficient (14.3) and height of microshoots (5.1 cm). The optimal concentration of auxin at the rooting stage has been established. The highest percentage of rooted microshoots (91%) was noted on a nutrient medium with 0.3 mg/l indoleacetic acid.

In Vitro Shoot Culture of Sesuvium portulacastrum: An Important Plant for Phytoremediation

Sesuvium portulacastrum L., a member of the family Aizoaceae, is an important coastal halophyte. Due to its adaptability to salinity and heavy metals, S. portulacastrum has now been widely used for the phytoremediation of saline soils and wastewater and the protection of the coast from erosion. The increasing use of this plant requires a large number of propagules. Stem cutting propagation and seed germination cannot meet this demand, and such propagations can initiate and spread diseases. A recent occurrence of Bipolaris sesuvii J.Z. Zhang and Gibbago trianthemae E.G. Simmons in S. portulacastrum resulted in the substantial loss of the plants during the remediation of aquaculture wastewater. Thus, there is an urgent need for establishing efficient methods of propagating disease-free starting materials. In the present study, we evaluated different growth regulators in the induction of axillary shoots from nodal explants cultured on Murashige and Skoog medium and identified that zeatin (ZT) and α-naphthaleneacetic acid (NAA) was an appropriate combination for inducing high numbers of axillary shoots. The nodal explants were then cultured on MS medium supplemented with different concentrations of ZT and NAA, and the combination of ZT at 1.0 mg L−1 and NAA at 0.3 mg L−1 induced more than 12 axillary shoots per explant. The axillary shoots were excised to produce microcuttings or microshoots, which were rooted on half-strength MS medium supplemented with different concentrations of indole-3-acetic acid (IAA) or indole-3-butyric acid (IBA). The results showed that IBA at 0.6 mg L−1 induced 91.7% of the microcuttings to root with root numbers of over 36 per cutting. The rooted plantlets were healthy and true-to-type and grew vigorously in plug trays or plastic containers with a 100% survey rate in a greenhouse. Thus, this established protocol could be used for the rapid propagation of genetically identical and disease-free plants of S. portulacastrum for phytoremediation and the protection of shoreline soils from erosion.

Propagation of Coccinia cordifolia (L.) Cogn. from shoot tip and nodal segment through micropropagation techniques

The present work was undertaken to develop a reproducible protocol for the micropropagation of an important medicinal plant of the Cucurbitaceae family, Coccinia cordifolia (L.) Cogn., by using shoot tips and nodal segments to overcome the impediment in seed settings and seed germination of conventional reproduction. To develop an efficient protocol, 0.1% HgCl2 treatment for 6 minutes was found effective for surface sterilization of field-grown explants to eliminate microbes and fungus and to get healthy tissues. Throughout the study, different concentrations of auxin, cytokine, and gibberellin were used either alone or in combination as supplemented in MS medium to find suitable conditions and suitable explant (nodal segment). BAP was the best cytokine source as 2.0 mg/l BAP produced 4.0±0.20 and 4.8±0.09 shoots per culture and gained 5.8±0.11 cm and 6.7±0.32 cm length with regeneration rate of 90 and 100 percent in shoot tips and node, respectively. The highest percentage (90%) of regeneration for axillary shoot proliferation was obtained in node explants at MS medium containing 2.0 mg/l BAP + 0.1 mg/l NAA. The highest number of shoots regenerated per culture was 3.0±0.41, with a length of 6.0±0.16 cm. GA3 was also found effective in producing longer shoots with the combination of BAP, but the average shoot number was reduced drastically. Although full-strength MS medium was found to be ideal for shoot regeneration and used in shoots proliferated experiments, half and full-strength of MS medium with auxins supplements of 0.5, 1.0, and 2.0 mg/l either of IBA, NAA, and IAA) were used for roots growth and half strength nutrients supplemented with 0.5 mg/l IBA was found most compelling. The highest number of roots regenerated per shoot was 3.1±0.30, and the average root length was 1.8±0.30 cm. Rooted plantlets were finally transplanted into small plastic pots containing sun sterilized sand, soil and humus (1: 2: 1) to adapt the plantlet in ex vitro environment, and acclimatized plantlets showed 95% survival rate in outdoor condition which proved the effectiveness of using biotechnology to improve plant’s growth rate and mass production.

Tissue-specific expression of barnase in tobacco delays axillary shoot development after topping.

Shoot branching is an important agronomic trait that determines plant architecture and affects crop productivity (Shen et al., 2019). Molecular signals from the shoot apical meristem (SAM) create a hormonal environment that integrates with the expression of axillary bud-specific repressors such as BRANCHED1 (BRC1) to inhibit axillary shoot formation (Wang et al., 2019 and references therein). The signal is eliminated by topping (SAM removal), enabling the formation of new shoots (suckers) from axillary buds (Figure 1a).

Micropropagation of Vaccinium virgatum ‘Delite’: a rabbiteye cultivar adapted to mild winters

ABSTRACT Blueberries have large amounts of antioxidant and anti-inflammatory compounds, playing an important role in improving health. In vitro culture is a possible method for vegetative propagation overcoming the problems of pathogen infection. Most studies on the in vitro culture of blueberries focus on highbush and lowbush cultivars. However, blueberry rabbiteye cultivars have lower chilling requirements and are more adapted to mild winter regions. Our objective was to develop a micropropagation protocol through axillary shoot growth, with reduced risks of somaclonal variation in ‘Delite’ rabbiteye blueberry. In vitro establishment was tested with single-node segments. The most promising results were using 2.5 µM zeatin. Multiplication through axillary shoot growth, starting with two-node stem segments were more successful in the concentration of 2.5 µM zeatin (70% of explants with axillary shoot growth). In vitro and ex vitro rooting with acclimatization were compared. In vitro rooting was achieved with immersion in 500 mg L−1 IBA (indole-3-butyric acid). Different substrates and mineral salts were also compared in ex vitro rooting. The results in vermiculite with WPM mineral salts were superior (68% rooting). In conclusion, this study provides techniques for micropropagation through axillary shoots of ‘Delite’ rabbiteye blueberry concerning establishment, multiplication, rooting, and acclimatization of plants.

Endophytic Bacteria in Banana In Vitro Cultures: Molecular Identification, Antibiotic Susceptibility, and Plant Survival

Microbial contamination is a common problem that causes significant losses in plant micropropagation systems. The present study reports on the identification and control of bacterial contaminants in banana in vitro cultures. Twelve isolates belonging to Bacillus pumilus (S2), Bacillus subtilis (R2 and M4), Geobacillus stearothermophilus (S1, S3, S4, P2, M3 and R3) and Paenibacillus spp. (P1, R1 and M2) were identified by sequencing of 16S rRNA, gyrA or gyrB genes. Antibiotic susceptibility testing was performed with the disk diffusion method on bacterial isolates using 36 antimicrobial agents. Some antibiotics, notably Ticarcillin, Penicillin, Ampicillin, Cefazolin and Imipenem, had a broader range of bactericidal activity than others did. When contaminated axillary shoot cultures of banana were treated with 100 or 200 mg·L−1 of ticarcillin, ampicillin or penicillin the bacteria were eliminated, but a reduction in shoot multiplication and growth was observed. These findings contribute to minimizing the losses in the commercial micropropagation of banana.

In vitro axillary shoot regeneration from cotyledonary node of Bishop’s weed (Trachyspermum ammi L.)

Bishop’s seed (Trachyspermum ammi L.) is a vital neutraceutial plant used as spice and also used as medicinal plant for curing different ailments and diseases of humans and animals. The study presents the in vitro regeneration of Bishop’s seed using cotyledonary node explant taken from in vitro germinated seeds. All mediums used in this study were comprised of MS (0.44%) medium having 3.0%sucrose and 0.65% agar with pH of 5.8. Expalnts were inoculated on MS medium enriched with 0.10-1.60 mg/L Thidiazuron (TDZ) alone or in combination with 0.10 mg/L IBA (Indole-3-butyric acid) for seven weeks under 16/8 light photoperiod provided with with cool-white fluorescent lamps. Thereafter, explants were sucultured on MS medium without any plant growth regulators (PGRs) for five weeks more under similar culture conditions. Results revealed 100% shoot regeneration frequency with 42.85-100.00%. The shoot count and shoot length ranged 3.43-19.40 and 1.04-2.74 cm respectively. The highest shoot count (19.40) and shoot length (2.74 cm) were observed on Ms medium enriched with 0.20 mg/L TDZ and 0.20 mg/L TDZ+0.10 mg/L IBA respectively. Relatively low rooting was recorded on IBA containing medium with low adaptation percentage of plantlets in pots containing peat moss.

Genome-wide identification of the tobacco GDSL family and apical meristem-specific expression conferred by the GDSL promoter

BackgroundGDSL esterases/lipases are a large protein subfamily defined by the distinct GDSL motif, and play important roles in plant development and stress responses. However, few studies have reported on the role of GDSLs in the growth and development of axillary buds. This work aims to identify the GDSL family members in tobacco and explore whether the NtGDSL gene contributes to development of the axillary bud in tobacco.ResultsOne hundred fifty-nine GDSL esterase/lipase genes from cultivated tobacco (Nicotiana tabacum) were identified, and the dynamic changes in the expression levels of 93 of these genes in response to topping, as assessed using transcriptome data of topping-induced axillary shoots, were analysed. In total, 13 GDSL esterase/lipase genes responded with changes in expression level. To identify genes and promoters that drive the tissue-specific expression in tobacco apical and axillary buds, the expression patterns of these 13 genes were verified using qRT-PCR. GUS activity and a lethal gene expression pattern driven by the NtGDSL127 promoter in transgenic tobacco demonstrated that NtGDSL127 is specifically expressed in apical buds, axillary buds, and flowers. Three separate deletions in the NtGDSL127 promoter demonstrated that a minimum upstream segment of 235 bp from the translation start site can drive the tissue-specific expression in the apical meristem. Additionally, NtGDSL127 responded to phytohormones, providing strategies for improving tobacco breeding and growth.ConclusionWe propose that in tobacco, the NtGDSL127 promoter directs expression specifically in the apical meristem and that expression is closely correlated with axillary bud development.

The Effect of Sucrose Supplementation on the Micropropagation of Salix viminalis L. Shoots in Semisolid Medium and Temporary Immersion Bioreactors

The effect of sucrose concentration on the micropropagation of axillary shoots of willow was investigated. The following factors were examined: the culture system (semisolid medium in glass jars versus liquid medium in temporary immersion bioreactors), the type of explant (apical and basal sections), the frequency of immersion, and CO2 enrichment. Shoots and leaf growth were significantly higher in RITA® bioreactors than in the jars for all the sucrose treatments. Apical or basal sections of willow cultured in bioreactors under high light intensity (150 µmol m−2 s−1) and ventilated six times a day with CO2-enriched air were successfully proliferated without sucrose, whereas shoots cultured in jars did not proliferate well if sucrose concentration was 0.5% or lower. More roots were formed when sucrose was added to the medium. Shoots cultured in bioreactors were successfully acclimatized irrespective of the sucrose treatment and the root biomass when transferred to ex vitro conditions. This is the first report of photoautotrophic willow micropropagation, our results confirm the importance of proper gaseous exchange to attain autotrophy during in vitro propagation.

Energy-Saving LED Light Affects the Efficiency of the Photosynthetic Apparatus and Carbohydrate Content in Gerbera jamesonii Bolus ex Hook. f. Axillary Shoots Multiplied In Vitro.

Simple SummaryGerbera is one of the most important ornamental plants on the cut flower market. The basic reproduction methods of numerous cultivars of this species are in vitro techniques. In in vitro cultures, all plant growth conditions are controlled, including the light (intensity, quality, and duration). In tissue cultures, light quality is the most important factor that influences plant morphogenesis (growth and development). Light emitting diodes (LEDs), in contrast to the commonly used fluorescent lamps, allow for adjusting the light quality to the specific requirements of plants. LEDs are also energy-efficient and contain no harmful substances (e.g., mercury). The aim of the study was to analyze the effect of different light qualities emitted by LEDs during in vitro multiplication of Gerbera on its metabolic and physiological development. We compared endogenous carbohydrate content in the tissues and the condition of the photosynthetic apparatus in plants grown under fluorescent lamps and LED light. The study showed that the use of LEDs did not disturb the secondary metabolism of carbohydrates and the multiplied shoots were of high quality. The mixture of red and blue LED light in a 7:3 proportion is recommended for gerbera micropropagation. This light quality positively influenced the functioning of the photosynthetic apparatus.An energy-saving light emitting diode (LED) system allows for adjustment of light quality, which affects plant development and metabolic processes in in vitro cultures. The study investigated the content of endogenous carbohydrates and the condition of the photosynthetic apparatus of Gerbera jamesonii Bolus ex Hook. f. Our aim was to analyze the effects of different LED light qualities—100% red light (R LED), 100% blue (B LED), a mixture of red and blue (7:3) (RB LED), and a fluorescent lamp as a control (Fl)—during the multiplication of axillary shoots. After 40 days, the culture measurements were performed using a non-invasive pulse amplitude modulation (PAM) fluorimeter. Sugar content was assessed with high performance liquid chromatography (HPLC). Two forms of free monosaccharides (glucose and fructose), two sugar alcohol derivatives (inositol and glycerol), and seven forms of free oligosaccharides were identified. Of those, glucose content was the highest. LEDs did not disturb the sugar metabolism in multiplied shoots. Their monosaccharides were three times more abundant than oligosaccharides; the same results were found in plants grown under control light. R light depleted the performance of the photosynthetic apparatus and caused its permanent damage. The RB LED spectrum ensured the most efficient non-photochemical quenching of the photosystem II (PS II) excitation state and high shoot quality.