Growth In Tissue Culture Research Articles

Optimization of crop tissue culture technology and its impact on biomolecular characteristics

Modern agriculture relies on crop tissue culture technology for fast propagation, genetic improvement, and protection of plant species. Conventional media like Murashige and Skoog (MS) usually lack optimal conditions for embryogenesis, necessitating the development of improved media tailored to specific crop requirements. In this article, we introduce an Efficient Grid Identified-Deep Feedforward Neurons (EGI-DFFN) to identify the ideal nutrient and vitamin levels of the crop plants for improving crop tissue culture aimed to improve crop plant growth in a lab setting by predicting callogenesis rate (CGR), embryogenesis rate (EGR), and somatic embryo number (SEN), shoot regeneration rate (SRR), rooting rate (RR).Different concentrations of ionic macronutrients, bio-molecular, and vitamins of the crop plant are the input to the predictive model, which is collected through the laboratory Callus Induction Experiment (CIE). Z-score normalization is used to preprocessing the CIE data to ensure consistent scales across different input features and improve model training performance. DFFN used discriminates to predict complex relationships and interactions between CGR, EGR, SEN, SRR, and RR with EGI tuning. The EGI-DFFN model has significantly improved crop tissue culture growth by accurately predicting the CGR, EGR, SEN, SRR, and RR respectively. The EGI-DFFN model enhances understanding of how ionic macronutrients and vitamins impact plant growth. It identifies optimal concentrations of the biomolecular to enhance somatic embryo formation and plantlet development, providing insights for optimizing crop tissue culture conditions for optimal growth outcomes.

Invasive Pulmonary Aspergillosis in Lung Transplant Recipients: Retrospective Clinical Analysis from a Tertiary Transplant Center.

Aspergillosis is the most common invasive fungal infection among lung transplant recipients (LTRs). Although its incidence is lower than that of bacterial or viral infections, it poses a similar or even higher mortality rate due to challenges in early diagnosis, limited treatment options, and various complications. Therefore, we aimed to evaluate the pulmonary aspergillosis cases in our tertiary lung transplant center. A retrospective analysis of 146 LTRs was performed. The demographic data, microbiological and histopathological test results, and radiological findings used for Aspergillus identification were recorded. Aspergillus spp. was detected in 13 of 146 LTRs (9%), mean age 42.5 ± 14.06 years, an average of 18.9 months after lung transplantation. 3 cases (23%) had Aspergillus growth in tissue culture, and 2 (15.4%) showed fungal elements with septal hyaline fibrils in tissue pathology. Aspergillus spp Polymerase chain reaction (PCR) was positive in bronchoalveolar lavage of 8 (61.5%) cases. In addition, 4 (30.7%) cases had relevant tomography findings. The most common pathogens were A. Terreus (21%), A. Fumigatus (14%), and A. Flavus (14%). The mortality rate was 15%. LTRs are at high risk of Aspergillus spp infections. Early diagnosis with microbiological, histopathological, and radiological tests, in addition to well-established prevention strategies, prophylaxis, and treatment will provide a better survival rate for patients.

Влияние 10-(6'-пластохинонил) децилтриметилфосфония (SkQ1) на образование каллусов и регенерацию растений, и их устойчивость к гипоксии

This work investigated the effect of 10-(6'-plastoquinonyl) decyltrimethylphosphonium (SkQ1) on the regeneration, growth and development of cell and tissue cultures of such plants as sugar cane (Saccharum officinarum L.), spring wheat (Triticum aestivum L.), corn (Zea mays L.), tobacco (Nicotiana tabacum L.), alfalfa of two species (Medicago glutinosa L.) and (Medicago falcata L.), cornflower (Thalictrum minus L.) and stephania (Stephania glabra Miers.). The stimulating effect of SkQ1 in concentrations from 0.5 to 10 nM has been shown, promoting better shoot and root formation in calli, an increase in growth rate, expressed in the cell mass of calli and their growth index. In addition, the effect of SkQ1 on the resistance of sugarcane calli to hypoxia was studied by assessing the growth index of calli of sensitive and hypoxia-resistant sugarcane lines when incubated under anaerobic conditions. It was shown that in both sensitive and resistant calli grown on a medium supplemented with 10 nM SkQ1, the growth index was 4 and 3.3 times higher, respectively, than in calli cultivated in a medium not containing SkQ1. Keywords: ANTIOXIDANTS, PLASTOQUINONE, 10-(6'-PLASTOQUINONYL) DECYLTRIMETHYLPHOSPHONIUM (SkQ1), CALLUS, EMPLANTS, REGENERATION, SUSPENSION CULTURE, HYPOXIA

Silver Nanoparticles Reduce the Toxic Effects of Cadmium on Datura stramonium Callus Culture

Cadmium (Cd) is one the most toxic metals harmful for both animals and plants. In this study, we test whether silver nanoparticles (AgNPs) can protect plants from cadmium toxicity. AgNPs treatments (0 and 100 μg/L) were applied to Datura stramonium calli grown in different cadmium metal environments (0, 150, 300, 450, and 600 μM). Cd application led to a decrease in fresh weight (FW), dry weight (DW), relative water content (RWC), total chlorophyll content (Chl.T), tolerance index (Ti), and bioaccumulation factor (BCF). The Cd treatment increased the hydrogen peroxide (H2O2), catalase (CAT), and guaiacol peroxidase (GPX) contents and Cd accumulation (Cd con). These results show the positive role of AgNPs in protecting the callus cultures from oxidative stress. The AgNPs pretreatment improved the growth of tissue cultures compared to nontreatment, increasing FW, DW, RWC, and Chl.T; the highest CAT and GPX activities were detected in the AgNPs pretreatment condition; and AgNPs pretreatment improved Ti and BCF, despite increased Cd. Also, this treatment caused a decrease in H2O2. Based on these results, we propose AgNPs as an effective agent to reduce the toxic effects of Cd metal on D. stramonium.

The Highly Conserved Stem-Loop II Motif Is Dispensable for SARS-CoV-2.

The stem-loop II motif (s2m) is an RNA structural element that is found in the 3' untranslated region (UTR) of many RNA viruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Though the motif was discovered over 25 years ago, its functional significance is unknown. In order to understand the importance of s2m, we created viruses with deletions or mutations of the s2m by reverse genetics and also evaluated a clinical isolate harboring a unique s2m deletion. Deletion or mutation of the s2m had no effect on growth in vitro or on growth and viral fitness in Syrian hamsters in vivo. We also compared the secondary structure of the 3' UTR of wild-type and s2m deletion viruses using selective 2'-hydroxyl acylation analyzed by primer extension and mutational profiling (SHAPE-MaP) and dimethyl sulfate mutational profiling and sequencing (DMS-MaPseq). These experiments demonstrate that the s2m forms an independent structure and that its deletion does not alter the overall remaining 3'-UTR RNA structure. Together, these findings suggest that s2m is dispensable for SARS-CoV-2. IMPORTANCE RNA viruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), contain functional structures to support virus replication, translation, and evasion of the host antiviral immune response. The 3' untranslated region of early isolates of SARS-CoV-2 contained a stem-loop II motif (s2m), which is an RNA structural element that is found in many RNA viruses. This motif was discovered over 25 years ago, but its functional significance is unknown. We created SARS-CoV-2 with deletions or mutations of the s2m and determined the effect of these changes on viral growth in tissue culture and in rodent models of infection. Deletion or mutation of the s2m element had no effect on growth in vitro or on growth and viral fitness in Syrian hamsters in vivo. We also observed no impact of the deletion on other known RNA structures in the same region of the genome. These experiments demonstrate that s2m is dispensable for SARS-CoV-2.

THE EFFECT OF SEVERAL CONCENTRATIONS OF GLYCINE AMINO ACID ADDITIONS IN MS MEDIUM TOWARS CALLUS GROWTH OF KONJAC (Amorphophallus muelleri) BY in vitro CULTURE

Konjac (Amorphophallus muelleri) is a plant that can be a source of carbohydrates and has high economic value. To accelerate the fulfillment of porang seeds in order to support maximum economic potential, it is necessary to develop alternative propagation techniques,which is in vitro culture. This study aimed to determine the effect of adding the glycine amino acid to MS medium on the growth of konjac callus and to determine the best concentration for callus growth in konjac plant tissue culture. This research was using frog bulb (bulbil) for the explant. In this study used a completely randomized design (CRD) with glycine amino acid concentrations from 0 mg/L, 1 mg/L, 2 mg/L, 3 mg/L, 4 mg/L, dan 5 mg/L with 3 replications. Data were analyzed using SPSS software with one way ANAVA and further test using DMRT 5%.The results showed that the addition of the glycine amino acid to MS medium affected the callus growth of konjac plant tissue culture based on the analysis of variance (ANAVA) test. The best concentration used based on the Duncan Multiple Range Test (DMRT) 5%, was 4 mg/L with the average callus appearance time of 3.06 wac and the average bending time of 2.1 wac.

Growth of Eucheuma cottonii seaweed from tissue culture with different time acclimatization in Hurun Bay

Constraints on the seaweed culture in Indonesia until now still rely on seeds obtained by vegetative propagation and have poor quality, which often causes a decrease in genetic variability. Planting seaweed seedlings from tissue cultures that have been acclimatized in a greenhouse laboratory is believed to improve the quality of the seaweed seeds. This research aimed to observe the growth of tissue culture acclimatized Eucheuma cottonii seaweed seedlings to different rearing times in greenhouses applied to floating raft cages. This study used a completely randomized design (CRD) with three treatments and three replications. Acclimatization treatment was conducted for 2 weeks, 4 weeks, and 6 weeks. Seaweed seedlings acclimatized in the greenhouse are reared on bamboo floating rafts in the sea, with an initial weight of 5 grams each. The length of maintenance at sea is 28 days. The resulting daily growth rate in two-week acclimatization was 2.92%, the four-week treatment was 2.93%, and the six-week treatment was 2.57%. in raising seedlings at sea, the highest daily growth rate value was in the four-week treatment of 7.25%, followed by the two-week treatment of 7.11% and the six-week treatment of 6.43%.

In vitro propagation and assessment of genetic stability in date palm as affected by chitosan and thidiazuron combinations

BackgroundMass propagation of date palm has attracted the interest of commercial producers. However, this technique still faces many obstacles that hinder production. This study investigated the effect of chitosan (CHT) at various concentrations for the possibility to apply it in combination with thidiazuron (TDZ) on the growth and development of tissue cultures of Barhee cultivar. ResultsThe results showed that CHT and TDZ on in vitro proliferation of Barhee date palm cultivar were significant. The highest response rate and the number of shoots per jar were found in MS media supplemented with 15 mgL–1 CHT and 0.5 mgL–1 TDZ combination. Furthermore, we found that the combined application between 20 mg L−1 CHT+ 1.0 mg L−1 TDZ resulted in the highest shoots content of endogenous IAA, compared with other treatments. At the same time, the data revealed that the maximum cytokinins (CKs) content of shoots occurred in a medium supplemented with 15 mg L−1 CHT and 0.5 mg L−1 TDZ. The genetic stability of the discussed micropropagation protocol was confirmed in this study by DNA-based technique RAPD (random amplified polymorphic DNA). The results may indicate that the micropropagation protocol developed in this research paper was appropriate and applicable for producing genetically stable date palm cv Barhee plants. ConclusionApplying the strategy of culture treatment with (CHT) and (TDZ) can be valuable for improving the propagation of date palm cv Barhee in vitro.

Economic Analysis of Micropropagation of Dragon Fruit (Hylocereus undatus (Haw.) Britton and Rose)

Cultivation of dragon fruit is rising in many topical and sub-tropical countries because of its high nutritional and medicinal values and is highly remunerative for the farmers. However, the supply of planting material is a bottleneck in meeting the increasing demand. With the upsurge in requirement for planting materials and the slow growth of the cactus, the conventional methods of stem cutting, which is the only source of acquiring planting materials at present, cannot meet the demand. Micropropagation which ensures the crop's clonal fidelity and availability of planting material throughout the year can be an alternative. Considering the higher cost of micropropagation, an economic analysis of in vitro regeneration for thousand plants was calculated and compared with the conventional stem cuttings. Cost of tissue cultured plantlets (Rs. 20.02) /plant) was found to be higher than the conventional method (Rs. 14.93 per plant). The major cost involved in micropropagation is attributed to the cost of skilled labours required for multiple sub-cultures, followed by the hardening media and tissue culture growth media. Nevertheless, the cost of tissue cultured plantlets is far lesser than the market price (Rs. 40/plant at university of Horticultural Sciences, Bagalkot). Hence, considering the clonality, uniformity and disease freeness of tissue cultured plantlets, micropropagation of dragon fruit may be considered feasible for both the producers and farmers. A further cost of production may be reduced with training skilled labours to improve their efficiency and other cheap sources of potting media may be explored.

Diyabetik Ayak Enfeksiyonu Olan Hastalarda Alt Ekstremite Damar Patolojilerinin İncelenmesi ve Klinik Sonuçları

Objective: Diabetes and vascular disorders raise the risk of diabetic foot infection and lower extremity amputation. Although DFI risk factors and microbiological analyses have been thoroughly researched, data for this specific group is limited. In this study, it was aimed to examine the underlying vascular risk factors of patients who were followed up and treated with the diagnosis of DFIs and clinical outcomes. Methods: Clinical, demographic, laboratory, microbiological, and foot examination data for 153 patients referred to our center for DFI between 2016 and 2021 were collected retrospectively from hospital information system. Results: The present center collected a total of 153 DFI patients during 5 year this period. There were 86 male and 67 women with a mean age of 67.71±15 years. There was 104 patients in non-vascular induced DFI group and 49 vascular induced DFI group. The top two comorbidities of DFI patients were cardiovascular disease and hypertension respectively. The rate of male patients was statistically higher in the vascular induced DFI group (p=0.003). Also hypertension and cardiovascular disease were more common in the vascular induced DFI group (p=0.0006, p=0.01). History of extremity amputation/debridement, having Wagner grade 5 DFI and Gram negative microorganism growth in tissue cultures were more common in the vascular induced DFI group (p=0.01, p=0.01 and p=0.0006). Extremity amputation/debridement rates were higher in the vascular induced DFI group (p=0.01) Conclusion: DFIs cause increased risk of amputation, prolonged antibiotic therapy, increased hospitalization, and increased healthcare costs as a result of investigations. Awareness of the vascular pathologies underlying DFIs can help clinicians manage the disease. The aim of this study is to emphasize the importance of vascular factors.

CO2 supplementation eliminates sugar-rich media requirement for plant propagation using a simple inexpensive temporary immersion photobioreactor

In vitro plant propagation systems such as temporary immersion bioreactors (TIBs) are valuable tools that enable production of disease-free plants with improved traits. However, TIB systems can be expensive, difficult to implement, and prone to contamination due to sugar rich propagation media. Using rapidly growing chicory root cultures to expedite design-build-test cycles, we report here an improved, low-cost version of a previously reported Hydrostatically-driven TIB (Hy-TIB) that facilitates economical use of gas mixtures. Bioreactor improvements include decreased material costs, expanded modes of operation, and a horizontal orientation of a plastic film plant growth chambers that increase propagule light exposure. To take advantage of these improvements, we describe here experiments that evaluate the impacts of elevated CO2 on propagation of cacao (Theobroma cacao) secondary embryos and nodal cultures of yam (Dioscorea spp.) during both phototrophic and photomixotrophic growth. Our experiments show that elevated CO2 during plant propagation significantly improved both cacao and yam propagule development and eliminated the need for supplemental sugars in tissue culture growth media. Thus, our improved Hy-TIB shows potential as a simple, low-cost, and scalable propagation platform with cost-effective gas composition control and reduced risk of contamination overgrowth. We provide detailed instructions for assembly of this Hy-TIB design and discuss the implications of its adoption in food-insecure regions of the world.

RGD-modified dextran hydrogel promotes follicle growth in three-dimensional ovarian tissue culture in mice

In vitro follicle growth is a promising technology to preserve fertility for cancer patients. We previously developed a three-dimensional (3-D) ovarian tissue culture system supported by mouse tumor cell-derived Matrigel. When murine ovarian tissues at 14 days old were cultured in Matrigel drops, antrum formation and oocyte competence were significantly enhanced compared with those cultured without Matrigel. In this study, we tested whether nonanimal-derived dextran hydrogels can support a 3-D ovarian tissue culture. We employed chemically defined dextran hydrogels consisting of dextran polymers crosslinked with polyethylene glycol (PEG)-based cell-degradable crosslinker. To determine the optimal gel elasticity for the 3-D tissue culture, we measured Young's modulus of dextran hydrogels at four concentrations (1.75, 2.25, 2.75, and 3.25 mmol/L), and cultured ovarian tissues in these gels for 7 days. As a result, 2.25 mmol/L dextran hydrogel with Young's modulus of 224 Pa was appropriate to provide physical support as well as to promote follicle expansion in the 3-D system. To mimic the natural extracellular matrix (ECM) environment, we modified the dextran hydrogels with two bioactive factors: ECM-derived Arg-Gly-Asp (RGD) peptides as a cell-adhesive factor, and activin A. The ovarian tissues were cultured in 2.25 mmol/L dextran hydrogels under four different conditions: Activin–/RGD– (A–R–), A + R–, A–R+, and A + R+. On Day 7 of culture, follicle and oocyte sizes were significantly increased in the RGD-modified conditions compared with those without RGD. The RGD-modified hydrogels also promoted mRNA levels of steroidogenic-related genes and estradiol production in the 3-D ovarian tissue culture. In vitro maturation and developmental competence of follicular oocytes were remarkably improved in the presence of RGD. In particular, blastocyst embryos were obtained only from A–R+ or A+R+ conditions after in vitro fertilization. We also determined synergistic effects of the RGD peptides and activin A on follicle growth and oocyte development in the 3-D tissue culture. In conclusion, our results suggest that RGD-modified dextran hydrogels provide an ECM-mimetic bioactive environment to support folliculogenesis in a 3-D ovarian tissue culture system.

Design of Centralized Intelligent Expert System and Contamination Detection of Tissue Cultured Sugarcane Crop

This paper presents the design a cloud based IoT enabled smart agriculture application for Hi-Tech tissue cultured sugarcane crop entitled “Design of Centralized Intelligent Expert System and Contamination Detection of Tissue Cultured Sugarcane Crop”. This expert system comprises of Raspberry Pi-4 (RPi), Arduino-Mega, GSM-Modem (Sim900) and sensor-modules for monitoring and control of essential parameters of laboratory for monitoring the physical parameters. The parameters monitored are temperature, humidity and light intensity of the tissue culture growth rooms with artificial day light timing and control, however, AI-based health prediction suggests the image processing for detection of culture contamination of sugarcane crop inside the growth-room. In addition, fire-smoke sensor and methane gas sensor are incorporated for fire protection and to avoid any disastrous situation. Three numbers of webcams are attached to the RPi for monitoring growth and health of explants. An AI-Model / weight was developed for detection of contamination that predicts the for health of Tissue Cultured Sugarcane Crop. Moreover, image enhancement was covered applying Generative Adversarial Networks (GAN)”. In this system, the RPi reads sensor's data through Arduino and convert it to data-frame with timestamp and geo-tag. The data along with the captured images are sent to a centralize cloud application for applying data mining and Artificial Intelligence; however, the model of contamination detection has been applied at edge device. This is to get meaningful insights of data for future decision making in maximizing crop yield and quality. Due to the great need of sugarcane crop in Pakistan, the Plant Tissue Culture (PTC) technology has been incorporated with Artificial Intelligence, the proposed system is aimed to be installed at established PTC-growth-rooms for sugarcane crop so the experts of field can be connected to the cloud application for its monitoring, control and data analytics. In addition, the use of telepresence through cloud application will enable PTC-experts to provide assistance to the remote user and resolve their issues timely, thus extending PTC technology all over the country which will eventually lead to increased crop yield with quality products in affordable price.

Influence of Substrate Composition and Container Size on the Growth of Tissue Culture Propagated Apple Rootstock Plants

Substrate composition and container size are considered crucial for apple rootstock grown in a plug seedling system. This study investigated the effect of substrate material’s proportion and different container volumes on the growth of apple rootstock (M-9) plants propagated by tissue culture. In substrate composition, three different ratios of peat moss (PM): vermiculite (VL): perlite (PL) at 1:1:1 (S1), 1:2:3 (S2), 3:1:2 (S3) were used. For container size, plants were grown in 1000 mL (C1), 500 mL (C2), and 300 mL (C3) containers filled by 1:1:1 mixture of PM, VL and PL. In both cases, plants were treated eleven weeks in a green house. Our results demonstrate that the plant height, number of leaves, leaf area, shoot fresh weight and root fresh weight of apple rootstock were significantly higher in substrate composition S3 compared with S1 composition. However, chlorophyll content (SPAD) and photosynthesis rate were unaffected by variation of substrate composition. Furthermore, rootstock grown in C1 container showed plant height, number of leaves, leaf area, number of nodes, root length, shoot and root’s fresh and dry weight were significantly higher than those grown in C2 and C3 volume containers. The chlorophyll content and transpiration rate were not significantly affected by the different container volumes. These results suggest that the substrate ratio 3:1:2 of PM:VL:PL and container size 1000 mL were more favorable than other treatments for initial growth and development of the tissue culture propagated apple rootstock plants.

Influence of light quality on growth and secondary metabolite in tissue cultures of Gardenia jasmonides, Variegata

The influence of white, blue and red Light-Emitting Diodes (LED) on the growth and the secondary metabolites production in calli and shoots cultures of Gardenia jasmonides, Variegata was evaluated. The white light generally has a powerful effect on both fresh and dry weights of the two types of cultures. White light treatment recorded (11.88 and 0.63 g) for the fresh and dry weight (in case of calli) followed by the blue light which recorded (11.78 and 0.60 g ), respectively. The extracts of shoots exposed to white LED light recorded the highest reducing efficiency (74.40 %). It could be justified as the shoots that exposed to the white light recorded the highest cholorogenic acid (CGA) compared to all the other treatments. Among different phenolic and flavonoids compounds which were tested in the calli and shoots cultures exposed to the different LED light quality, only the chlorogenic acid was detected in the different treatments in both cultures with varying quantities. The white LED light recorded the highest content of chlorogenic acid with both cultures, since it scored 52.43 µg / gdw with calli cultures versus 8783.53 µg / gdw with shoots cultures.

Vaccinia Virus Expressing Interferon Regulatory Factor 3 Induces Higher Protective Immune Responses against Lethal Poxvirus Challenge in Atopic Organism.

Vaccinia virus (VACV) is an enveloped DNA virus from the Orthopoxvirus family, various strains of which were used in the successful eradication campaign against smallpox. Both original and newer VACV-based replicating vaccines reveal a risk of serious complications in atopic individuals. VACV encodes various factors interfering with host immune responses at multiple levels. In atopic skin, the production of type I interferon is compromised, while VACV specifically inhibits the phosphorylation of the Interferon Regulatory Factor 3 (IRF-3) and expression of interferons. To overcome this block, we generated a recombinant VACV-expressing murine IRF-3 (WR-IRF3) and characterized its effects on virus growth, cytokine expression and apoptosis in tissue cultures and in spontaneously atopic Nc/Nga and control Balb/c mice. Further, we explored the induction of protective immune responses against a lethal dose of wild-type WR, the surrogate of smallpox. We demonstrate that the overexpression of IRF-3 by WR-IRF3 increases the expression of type I interferon, modulates the expression of several cytokines and induces superior protective immune responses against a lethal poxvirus challenge in both Nc/Nga and Balb/c mice. Additionally, the results may be informative for design of other virus-based vaccines or for therapy of different viral infections.

Variables Affecting Shoot Growth and Plantlet Recovery in Tissue Cultures of Drug-Type Cannabis sativa L.

Tissue culture approaches are widely used in crop plants for the purposes of micropropagation, regeneration of plants through organogenesis, obtaining pathogen-free plantlets from meristem culture, and developing genetically modified plants. In this research, we evaluated variables that can influence the success of shoot growth and plantlet production in tissue cultures of drug-type Cannabis sativa L. (marijuana). Various sterilization methods were tested to ensure shoot development from nodal explants by limiting the frequency of contaminating endophytes, which otherwise caused the death of explants. Seven commercially grown tetrahydrocannabinol (THC)-containing cannabis genotypes (strains) showed significant differences in response to shoot growth from meristems and nodal explants on Murashige and Skoog (MS) medium containing thidiazuron (1 μM) and naphthaleneacetic acid (0.5 μM) plus 1% activated charcoal. The effect of Driver and Kuniyuki Walnut (DKW) or MS basal salts in media on shoot length and leaf numbers from nodal explants was compared and showed genotype dependency with regard to the growth response. To obtain rooted plantlets, shoots from meristems and nodal explants of genotype Moby Dick were evaluated for rooting, following the addition of sodium metasilicate, silver nitrate, indole-3-butyric acid (IBA), kinetin, or 2,4-D. Sodium metasilicate improved the visual appearance of the foliage and improved the rate of rooting. Silver nitrate also promoted rooting. Following acclimatization, plantlet survival in hydroponic culture, peat plugs, and rockwool substrate was 57, 76, and 83%, respectively. The development of plantlets from meristems is described for the first time in C. sativa and has potential for obtaining pathogen-free plants. The callogenesis response of leaf explants of 11 genotypes on MS medium without activated charcoal was 35% to 100%, depending on the genotype; organogenesis was not observed. The success in recovery of plantlets from meristems and nodal explants is influenced by cannabis genotype, degree of endophytic contamination of the explants, and frequency of rooting. The procedures described here have potential applications for research and commercial utility to obtain plantlets in stage 1 tissue cultures of C. sativa.

Stimulation of Nicotiana tabacum L. In Vitro Shoot Growth by Endophytic Bacillus cereus Group Bacteria.

In vitro plant tissue cultures face various unfavorable conditions, such as mechanical damage, osmotic shock, and phytohormone imbalance, which can be detrimental to culture viability, growth efficiency, and genetic stability. Recent studies have revealed a presence of diverse endophytic bacteria, suggesting that engineering of the endophytic microbiome of in vitro plant tissues has the potential to improve their acclimatization and growth. Therefore, the aim of this study was to identify cultivated tobacco (Nicotiana tabacum L.) endophytic bacteria isolates that are capable of promoting the biomass accumulation of in vitro tobacco shoots. Forty-five endophytic bacteria isolates were obtained from greenhouse-grown tobacco plant leaves and were assigned to seven Bacillus spp. and one Pseudomonas sp. based on 16S rRNA or genome sequence data. To evaluate the bacterial effect on in vitro plant growth, tobacco shoots were inoculated with 22 isolates selected from distinct taxonomic groups. Four isolates of Bacillus cereus group species B. toyonensis, B. wiedmannii and B. mycoides promoted shoot growth by 11–21%. Furthermore, a contrasting effect on shoot growth was found among several isolates of the same species, suggesting the presence of strain-specific interaction with the plant host. Comparative analysis of genome assemblies was performed on the two closely related B. toyonensis isolates with contrasting plant growth-modulating properties. This revealed distinct structures of the genomic regions, including a putative enzyme cluster involved in the biosynthesis of linear azol(in)e-containing peptides and polysaccharides. However, the function of these clusters and their significance in plant-promoting activity remains elusive, and the observed contrasting effects on shoot growth are more likely to result from genomic sequence variations leading to differences in metabolic or gene expression activity. The Bacillus spp. isolates with shoot-growth-promoting properties have a potential application in improving the growth of plant tissue cultures in vitro.

In vitro plant regeneration system for date palm (Phoenix dactylifera L.): effect of chelated iron sources

BackgroundIron chelate sources and their concentrations are important factors in in vitro propagation of date palm. This study’s objective was to investigate the effect of the iron chelated form on the growth and development of tissue cultures of Barhee cultivar. ResultsThe addition of FeEDDHA to the culture medium was more effective than FeEDTA on callus growth, shoot regeneration, and the number of shoots per jar, where the best result (220.8mg callus, 86.67% and 17.2 shoots per jar, respectively) was obtained by using 93.5 mg L−1 FeEDDHA (5.6 mg L−1 Fe), compared with other treatments. The results also indicate that using 93.5 mg L−1 FeEDDHA (5.6 mg L−1 Fe) as a supplement can decrease antioxidant enzymes CAT and POD activity compared to the rest of the treatments. Medium equipped with 187.0 mg L−1 FeEDDHA (11.2 mg L−1Fe) had the highest rooting percentage and number of roots per shoot than other treatments. The biochemical analysis results showed that treatments with FeEDDHA of 280.5 mg L−1 (16.8 mg L−1 Fe) and 187.0 mg L−1 (11.2 mg L−1Fe) significantly increased the iron content. The results showed that shoot maximum chlorophyll and endogenous IAA level content were recorded in a medium supplemented with 187.0 mg L−1 FeEDDHA (11.2 mg L−1Fe) as Fe source. ConclusionFeEDDHA used in the present study was proven to be a promising iron chelate source in comparison with the FeEDTA sources.

Development of an efficient protocol for production of healthy sugarcane seed cane through Meristem culture

Healthy seeds are a prerequisite for improving the productivity of sugarcane crop. The best way to ensure the production of disease-free sugarcane seeds is via a technique called ‘Meristem Culture’ (Tissue Culture). An efficient protocol for meristem culture of sugarcane has been developed in this study. The potting mix for the survival of meristem cultured plantlets in a greenhouse was standardized, and agronomy for growing tissue culture plantlets in the field was developed. Multiple shoots were observed on Murashige and Skoog's (MS) medium supplemented with BAP 0.2 ​mg/l and Kinetin 0.1 ​mg/l. Best rooting was observed on MS media supplemented with IAA 1 ​mg/l and NAA 1 ​mg/l. Soil mixture containing [Soil ​+ ​Vermiculite ​+ ​Sand in 4: 1: 1 proportion (by volume)] ​+ ​Vermicompost @ 25% by volume of medium ​+ ​NPK is best suited for hardening tissue cultured plants in a greenhouse. Two feet distance between 2 plants and 3 feet distance between 2 rows was found as an appropriate method for planting tissue cultured plantlets in the field. Setts obtained from tissue cultured plants planted at 15 ​cm spacing gave higher yield than the other spacing. Setts obtained from micro-propagated plants (M) gave the highest yield as compared to setts from conventional sugarcane raising. The protocol developed in this study, shows potential for development of sugarcane meristem cultured plantlets and use of these plants for planting in field to obtain healthy sugarcane seed.