Tissue Culture System Research Articles

An Efficient Method for the Propagation of Bougainvillea glabra ‘New River’ (Nyctaginaceae) from In Vitro Stem Segments

Bougainvillea, an evergreen climbing shrub of the Nyctaginaceae family, holds significant ornamental, economic, and medicinal value. Bougainvillea glabra ‘New River’ is widely used in landscapes due to its strong adaptability to the environment, abundance of flowers, and frequent flowering. Traditionally, Bougainvillea glabra ‘New River’ cultivation has relied on methods such as cuttings or grafting, with limited research on in vitro tissue culture propagation. This study aimed to optimize the tissue culture system, exploring a combination of plant growth regulators (PGRs) for Bougainvillea regeneration from in vitro stem segments. The Murashige and Skoog (MS) medium supplemented with indole-3-butyric acid (IBA), 6-benzylaminopurine (6-BA), and 1-naphthlcetic acid (NAA) was employed. The optimal sterilization of Bougainvillea stem segments involved a 30 s treatment with 75% alcohol and 10 min with 1% NaClO. The synergistic effect of 0.1 mg·L−1 of NAA and 2.5 mg·L−1 of 6-BA maximized the shoot sprouting frequency, while 2.5 mg·L−1 of 6-BA and 0.1 mg·L−1 of NAA produced the maximum shoots. Furthermore, 1.5 mg·L−1 of IBA and 0.1 mg·L−1 of NAA induced the highest rooting levels. This work demonstrates the successful adaptation of a greenhouse environment to efficiently regenerate plants in vitro from stem segments. This approach allows for the mass production of Bougainvillea glabra ‘New River’.

Lung Organoids: Systematic Review of Recent Advancements and its Future Perspectives.

Organoids are essentially an in vitro (lab-grown) three-dimensional tissue culture system model that meticulously replicates the structure and physiology of human organs. A few of the present applications of organoids are in the basic biological research area, molecular medicine and pharmaceutical drug testing. Organoids are crucial in connecting the gap between animal models and human clinical trials during the drug discovery process, which significantly lowers the time duration and cost associated with each stage of testing. Likewise, they can be used to understand cell-to-cell interactions, a crucial aspect of tissue biology and regeneration, and to model disease pathogenesis at various stages of thedisease. Lung organoids can be utilized to explore numerous pathophysiological activities of a lung since they share similarities with its function. Researchers have been trying to recreate the complex nature of the lung by developing various "Lung organoids" models.This article is a systematic review of various developments of lung organoids and their potential progenitors. It also covers the in-depth applications of lung organoids for the advancement of translational research. The review discusses the methodologies to establish different types of lung organoids for studying the regenerative capability of the respiratory system and comprehending various respiratory diseases.Respiratory diseases are among the most common worldwide, and the growing burden must be addressed instantaneously.Lung organoids along with diverse bio-engineering tools and technologies will serve as a novel model for studying thepathophysiology of various respiratory diseasesand fordrug screening purposes.

The Impacts of Plant Growth Regulators on the Rapid Propagation of Gardenia jasminoides Ellis. in Tissue Culture

The optimization of explant selection and adjustment of plant growth regulators (PGRs) ratio may enhance the efficiency of micro-propagation of Gardenia jasminoides Ellis. The findings of the study suggest that the shoot tip proved to be the optimal explant for regenerating adventitious buds, with an impressive regeneration rate of 77.78% and the average number of adventitious buds being 2.86. The ideal medium consisted of Murashige and Skoog (MS) medium supplemented with 6-benzylaminopurine (6-BA) at a 2 mg L−1, indoleacetic acid (IAA) at a 0.2 mg L−1, kinetin (KT) at 0.15 mg L−1, resulting in an outstanding regeneration rate of adventitious buds reaching up to 91.11%. For rooting purposes, the best medium was found be half-strength MS supplemented with indoleacetic acid (IAA) 0.5 mg L−1, achieving an rate for adventitious roots amounting to as high as 97.78%. The culture plantlets ultimately thrived, achieving an impressive transplanting survival rate of 93.33%. The application of PGRs was also found to enhance the regeneration of adventitious buds by increasing the ratios of endogenous hormones ZR/IAA and GA3/IAA. Additionally, it facilitated the differentiation of adventitious roots by elevating the ratios of endogenous hormones IAA/ZR, IAA/GA3, and ABA/GA3. Our study would provide a theoretical reference for the establishment of an efficient gardenia tissue culture system and the industrial production of gardenia.

Injection-based hairy root induction and plant regeneration techniques in Brassicaceae

BackgroundHairy roots constitute a valuable tissue culture system for species that are difficult to propagate through conventional seed-based methods. Moreover, the generation of transgenic plants derived from hairy roots can be facilitated by employing carefully designed hormone-containing media.ResultsWe initiated hairy root formation in the rare crucifer species Asperuginoides axillaris via an injection-based protocol using the Agrobacterium strain C58C1 harboring a hairy root-inducing (Ri) plasmid and successfully regenerated plants from established hairy root lines. Our study confirms the genetic stability of both hairy roots and their derived regenerants and highlights their utility as a permanent source of mitotic chromosomes for cytogenetic investigations. Additionally, we have developed an effective embryo rescue protocol to circumvent seed dormancy issues in A. axillaris seeds. By using inflorescence primary stems of Arabidopsis thaliana and Cardamine hirsuta as starting material, we also established hairy root lines that were subsequently used for regeneration studies.ConclusionWe developed efficient hairy root transformation and regeneration protocols for various crucifers, namely A. axillaris, A. thaliana, and C. hirsuta. Hairy roots and derived regenerants can serve as a continuous source of plant material for molecular and cytogenetic analyses.

FGF-stimulated tendon cells embrace a chondrogenic fate with BMP7 in newt tissue culture.

Newts can regenerate functional elbow joints after amputation at the joint level. Previous studies have suggested the potential contribution of cells from residual tendon tissues to joint cartilage regeneration. A serum-free tissue culture system for tendons was established to explore cell dynamics during joint regeneration. Culturing isolated tendons in this system, stimulated by regeneration-related factors, such as fibroblast growth factor (FGF) and platelet-derived growth factor, led to robust cell migration and proliferation. Moreover, cells proliferating in an FGF-rich environment differentiated into Sox9-positive chondrocytes upon BMP7 introduction. These findings suggest that FGF-stimulated cells from tendons may aid in joint cartilage regeneration during functional elbow joint regeneration in newts.

Rapid prototyping of PMMA-based microfluidic spheroid-on-a-chip models using micromilling and vapour-assisted thermal bonding

The application of microfluidic devices as next-generation cell and tissue culture systems has increased impressively in the last decades. With that, a plethora of materials as well as fabrication methods for these devices have emerged. Here, we describe the rapid prototyping of microfluidic devices, using micromilling and vapour-assisted thermal bonding of polymethyl methacrylate (PMMA), to create a spheroid-on-a-chip culture system. Surface roughness of the micromilled structures was assessed using scanning electron microscopy (SEM) and atomic force microscopy (AFM), showing that the fabrication procedure can impact the surface quality of micromilled substrates with milling tracks that can be readily observed in micromilled channels. A roughness of approximately 153 nm was created. Chloroform vapour-assisted bonding was used for simultaneous surface smoothing and bonding. A 30-s treatment with chloroform-vapour was able to reduce the surface roughness and smooth it to approximately 39 nm roughness. Subsequent bonding of multilayer PMMA-based microfluidic chips created a durable assembly, as shown by tensile testing. MDA-MB-231 breast cancer cells were cultured as multicellular tumour spheroids in the device and their characteristics evaluated using immunofluorescence staining. Spheroids could be successfully maintained for at least three weeks. They consisted of a characteristic hypoxic core, along with expression of the quiescence marker, p27kip1. This core was surrounded by a ring of Ki67-positive, proliferative cells. Overall, the method described represents a versatile approach to generate microfluidic devices compatible with biological applications.

Towards the fourth decade of IOPRI’s oil palm clones: Upcoming new variety

Experience of IOPRI on oil palm tissue culture would be four decades in 2024. The first decade was theperiod of oil palm clone development and field testing. The second decade was a setback for oil palm research because of a high percentage of oil palm clone abnormalities in the field. Moving on from these experiences, tissueculture systems such as: including improving the protocol, controlling the duration of callus induction, delimitating the number of subcultures, and building of oil palm clones database, were managed to be improved. In this way, alow abnormality level is under control and each clone is traceable. It is also noted that the oil palm clone provides20-30% higher production than that derived from DxP crossing seed due to the more uniform growth of generatedplants. Besides the protocol, ortet selection is also key to improvement. Ortets are elite plants from selected progeny testing or re-clone of the progeny testing. Molecular analysis is applied as a control of genetic variation possibility, which may occur in a particular phase due to specific stimuli. However, until the present time, a clonevariety has not been released yet. Therefore, in the fourth decade, mass production is expected and oil palm clonevarieties will officially be released and commercialized

Selective Targeting of Tumor Cells in a Microfluidic Tumor Model with Multiple Cell Types.

Organ-on-a-chip technology allows researchers to precisely monitor drug efficacy in 3D tissue culture systems that are physiologically more relevant to humans compared to 2D cultures and that allow better control over experimental conditions as compared to animal models. Specifically, the high control over microenvironmental conditions combined with the broad range of direct measurements that can be performed in these systems makes organ-on-a-chip devices a versatile tool to investigate tumor targeting and drug delivery. Here, we describe a detailed protocol for studying the cell-selective targeting of protein drugs to tumor cells on an organ-on-a-chip system using a co-culture consisting of BT-474 cancer cells and C5120 human fibroblasts as an example.

Micropropagation of Duboisia Species via Shoot Tip Meristem

Duboisia is an Australian native, commercially valuable for tropane alkaloid extraction. Clonal propagation of elite selections is essential to establish highly productive plantations. The current propagation system using stem cuttings is proven to be inefficient, prompting the industry to seek a more efficient and effective propagation tool. Tissue culture is a cost-effective alternative for mass propagation of true-to-type plants, particularly ideal for propagating elite Duboisia selections. In this context, attempts were made to develop a commercially viable high throughput micropropagation system for three Duboisia species: Duboisia myoporoides, Duboisia leichhradtii and Duboisia hopwoodii. Various nutrient media, hormone combinations and incubating conditions were tested to optimise each stage of the micropropagation pipeline. The findings revealed that the tissue culture media composition and hormone requirements are species-specific. With the optimised conditions, an efficient tissue culture system was developed, achieving successful meristem induction and multiplication. Species-specific rooting protocol optimisation resulted in 100% rooting for D. myoporoides and D. leichhardtii, and 70% rooting for D. hopwoodii. Furthermore, an optimised acclimatisation protocol supported 100% survival of D. myoporoides and D. leichhardtii and 80% of D. hopwoodii plantlets. This study, for the first time, demonstrated the capacity of successful meristem culture of three Duboisia species, establishing the foundation for high throughput micropropagation of Duboisia species.

Molecular symmetry breaking in the Frizzled-dependent planar polarity pathway

The core planar polarity pathway consists of six proteins that form asymmetric intercellular complexes that segregate to opposite cell ends in developing tissues and specify polarized cell structures or behaviors. Within these complexes, the atypical cadherin Flamingo localizes on both sides of intercellular junctions, where it interacts homophilically in trans via its cadherin repeats, whereas the transmembrane proteins Frizzled and Strabismus localize to the opposite sides of apposing junctions. However, the molecular mechanisms underlying the formation of such asymmetric complexes are poorly understood. Using a novel tissue culture system, we determine the minimum requirements for asymmetric complex assembly in the absence of confounding feedback mechanisms. We show that complexes are intrinsically asymmetric and that an interaction of Frizzled and Flamingo in one cell with Flamingo in the neighboring cell is the key symmetry-breaking step. In contrast, Strabismus is unable to promote homophilic Flamingo trans binding and is only recruited into complexes once Frizzled has entered on the opposite side. This interaction with Strabismus requires intact intracellular loops of the seven-pass transmembrane domain of Flamingo. Once recruited, Strabismus stabilizes the intercellular complexes together with the three cytoplasmic core proteins. We propose a model whereby Flamingo exists in a closed conformation and binding of Frizzled in one cell results in a conformational change that allows its cadherin repeats to interact with a Flamingo molecule in the neighboring cell. Flamingo in the adjacent cell then undergoes a further change in the seven-pass transmembrane region that promotes the recruitment of Strabismus.

Metabolic reprogramming driven by EZH2 inhibition depends on cell–matrix interactions

EZH2 (Enhancer of Zeste Homolog 2), a subunit of Polycomb Repressive Complex 2 (PRC2), catalyzes the trimethylation of histone H3 at lysine 27 (H3K27me3), which represses expression of genes. It also has PRC2-independent functions, including transcriptional coactivation of oncogenes, and is frequently overexpressed in lung cancers. Clinically, EZH2 inhibition can be achieved with the FDA-approved drug EPZ-6438 (tazemetostat). To realize the full potential of EZH2 blockade, it is critical to understand how cell-cell/cell-matrix interactions present in three-dimensional (3D) tissue and cell culture systems influences this blockade in terms of growth-related metabolic functions. Here, we show that EZH2 suppression reduced growth of human lung adenocarcinoma A549 cells in two-dimensional (2D) cultures but stimulated growth in 3D culture. To understand the metabolic underpinnings, we employed [13C6]-glucose Stable Isotope-Resolved Metabolomics (SIRM) to determine the effect of EZH2 suppression on metabolic networks in 2D versus 3D A549 cultures. The Krebs cycle, neoribogenesis, γ-aminobutyrate (GAB) metabolism, and salvage synthesis of purine nucleotides were activated by EZH2 suppression in 3D spheroids but not in 2D cells, consistent with the growth effect. Using simultaneous 2H7-glucose + 13C5,15N2-Gln tracers and EPZ-6438 inhibition of H3 trimethylation, we delineated the effects on the Krebs cycle, γ-aminobutyrate metabolism, gluconeogenesis, and purine salvage to be PRC2 dependent. Furthermore, the growth/metabolic effects differed for mouse Matrigel versus self-produced A549 extracellular matrix. Thus, our findings highlight the importance of the presence and nature of extracellular matrix in studying the function of EZH2 and its inhibitors in cancer cells for modeling the in vivo outcomes.

The effect of plant growth regulators, carbon sources and gelling agent on the callus regeneration of Malaysian rice MR219 (Oryza sativa L.)

The development of a highly efficient tissue culture system for indica rice is crucial to speed up the implementation of gene editing in breeding programmes as well as to explore the roles of indica genes. However, because indica rice types are recalcitrant to in vitro response, there are only a few reports on the successful transformation process on indica rice due to recalcitrant response to in vitro response. The present study attempted to identify the effects of plant growth regulator (PGR), carbon sources and different concentrations of gelling agent on embryogenic callus induction and rice regeneration of Oryza sativa var MR219. The PGR used were 1 or 2 mg/L 6-benzylaminopurine (BAP) and 1-Naphthaleneacetic acid (NAA) and the carbon sources were 30 g/L sucrose and maltose. Three different gelling agent concentrations (3 mg/L, 6 mg/L or 9 mg/L) were evaluated. The highest frequency of friable callus induction (87%) was observed in callus induction media (CI) containing Murashige and Skoong media (MS) supplemented with 3 mg/L 2,4-dichlorophenoxy acetic acid (2,4-D) and 0.3 mg/L kinetin. Histological studies of the calli showed the presence of embryogenic calli with small intercellular spaces compared to non-embryogenic areas which developed on the hormone MS media. The highest regeneration frequency (50%) was achieved when friable calli were cultured on MS medium containing 3% sucrose, 1 mg/L BAP, and 1 mg/L NAA with 9% phytagel. The callus grown on sucrose media showed the highest number (66.13%) of albino plantlets, while the number of albino plantlets on maltose media was smaller (20%). In conclusion, this optimised mature embryo-derived callus of MR219 rice is responsive to regeneration and could be amenable to genetic transformation studies.

Modern Approaches to In Vitro Clonal Banana Production: Next-Generation Tissue Culture Systems

In this study, the solid culture method, and Plantform™ and SETIS™ temporary immersion bioreactor systems were used comparatively to propagate, root, and acclimatize ‘Grande Naine’ and ‘Azman’ banana varieties for rapid, cheap, and mass production in in vitro conditions. Micropropagation rate, plant height, number of leaves, and fresh and dry weight parameters were investigated in the micropropagation stage across eight subcultures. Rooting rate, plant height, number of leaves, number of roots/plant, root length, fresh and dry weight parameters were investigated in the rooting stage. Photosynthetic pigment analyses and stoma examinations were performed throughout all stages. In the micropropagation stage, a 20% increase in the Plantform™ system, a 12% increase in the SETIS™ system in ‘Grande Naine’, an 82% increase in the Plantform™ system, and a 98% increase in SETIS™ system in ‘Azman’ were determined compared to the solid culture. At the rooting stage, higher data were obtained from bioreactor systems than solid culture. Plants from bioreactor systems acclimatized faster and developed healthier in the greenhouse stage. It was determined that stomata were more active, and pigment accumulation was higher in bioreactor systems. Genetic variations across subcultures are among the most critical issues in banana clonal propagation. Leaf samples were taken from each system, and plant variation was investigated using SSR (Simple Sequence Repeat) markers. No variation was observed from the initial stage to the greenhouse stage. As a result, it has been determined that bioreactor systems are an essential alternative for the mass production of bananas.

Shoot Organogenesis and Regeneration from Leaf Seedlings of Diospyros oleifera Cheng.

Persimmons (Diospyros) are economically important trees that are widely cultivated for wood production in China, and Diospyros oleifera Cheng is the main persimmon grafting stock. However, an efficient tissue culture system has not been perfected for D. oleifera due to the limits of proliferation and rooting cultures. Therefore, this study examined the effects of different plant growth regulators and concentrations on the primary culture of young embryos, induction of leaf callus, differentiation of adventitious shoots, and rooting culture of D. oleifera. The optimal formula for young embryo germination was 1/2 Murashige and Skoog (MS) medium containing 0.5 mg/L gibberellic acid (GA3); after 25 days, the sprouting rate of the young embryos was 67.3%. The best medium for leaf callus induction was 1/2MS medium containing 2.0 mg/L of 6-benzylaminopurine (6-BA) and 0.5 mg/L of naphthaleneacetic acid (NAA), and the callus induction rate was 88.9%. Then, the callus was transferred to 1/2MS medium containing 2.0 mg/L of zeatin (ZT), 0.5 mg/L of NAA, and 2.0 mg/L of thidiazuron (TDZ) to induce adventitious shoots; after 25 days, 5.4 buds were produced per explant, and the induction rate of the adventitious shoots was 88.3%. The adventitious shoots were transferred to 1/2MS medium containing 2.0 mg/L of ZT, 2.0 mg/L of 6-(γ,γ-dimethylallylamino)purine (2iP), and 0.1 mg/L of indole acetic acid (IAA) for the proliferation culture, for which the multiplication coefficient approached 7.5. After multiplication, the adventitious shoots were inoculated into 1/2MS medium containing 1.0 mg/L of indole butyric acid (IBA), 0.5 mg/L of NAA, and 1.0 mg/L of kinetin (KT); the rooting rate was 60.2%, and the average number of roots was 6.9.

In vitro and ex vitro propagation of Turkish myrtles through conventional and plantform bioreactor systems.

The myrtle (Myrtus communis) plant naturally grows in the temperate Mediterranean and subtropical regions and is used for various purposes; thus, it is among the promising species of horticultural crops. This study aimed to evaluate and compare the performance of different propagation systems, including rooting, solid media propagation, rooting, and with the Plantform bioreactor system, in achieving healthy and rapid growth of four myrtle genotypes with diverse genetic origins and well-regional adaptation. The selection of myrtle genotypes with distinct genetic backgrounds and proven adaptability to specific regions allowed for a comprehensive assessment of the propagation systems under investigation. Present findings proved that the Plantform system, the new-generation tissue culture system, was quite successful in micropropagation and rooting myrtle genotypes. We succeeded in vitro micropropagation and rooting of diverse wild myrtle genotypes, enabling year-round propagation without reliance on specific seasons or environmental conditions. The process involved initiating cultures from explants and multiplying them through shoot proliferation in a controlled environment. This contributes to sustainable plant propagation, preserving and utilizing genetic resources for conservation and agriculture.

Establishment of a high-efficiency transformation and genome editing method for an essential vegetable and medicine Solanum nigrum.

Solanum nigrum, which belongs to the Solanaceae family, is an essential plant for food and medicine. It has many important secondary compounds, including glycoproteins, glycoalkaloids, polyphenolics, and anthocyanin-rich purple berries, as well as many ideal characteristics such as self-fertilization, a short life cycle and a small genome size that make it a potential model plant for the study of secondary metabolism and fruit development. In this study, we report a highly efficient and convenient tissue culture, transformation and genome editing method for S. nigrum using leaf segments after 8 weeks of tissue culture, with a required period from transformation initiation to harvest of about 3.5 months. Our results also show multi-shoot regeneration per leaf segment and a 100% shoot regeneration efficiency in a shoot regeneration medium. Moreover, over 82% of kanamycin-resistant plants exhibited strong green fluorescence marker protein expression, with genetic integration confirmed by PCR results and green fluorescence protein expression in their T1 progeny. Furthermore, we successfully applied this transformation method to achieve an average of 83% genome editing efficiency of SnMYB1, a gene involved in regulating the anthocyanin biosynthetic pathway of S. nigrum in response to missing nutrients. Taken together, the combination of highly efficient tissue culture, transformation and genome editing systems can provide a powerful platform for supporting fundamental research on the molecular mechanisms of secondary metabolism, fruit development, and production of important compounds by biotechnology.

Exploring Genetic and Epigenetic Changes in Lingonberry Using Molecular Markers: Implications for Clonal Propagation.

Lingonberry (Vaccinium vitis-idaea L.) is an important and valuable horticultural crop due to its high antioxidant properties. Plant tissue culture is an advanced propagation system employed in horticultural crops. However, the progeny derived using this technique may not be true-to-type. In order to obtain the maximum return of any agricultural enterprise, uniformity of planting materials is necessary, which sometimes is not achieved due to genetic and epigenetic instabilities under in vitro culture. Therefore, we analyzed morphological traits and genetic and epigenetic variations under tissue-culture and greenhouse conditions in lingonberry using molecular markers. Leaf length and leaf width under greenhouse conditions and shoot number per explant, shoot height and shoot vigor under in vitro conditions were higher in hybrid H1 compared to the cultivar Erntedank. Clonal fidelity study using one expressed sequence tag (EST)-polymerase chain reaction (PCR), five EST-simple sequence repeat (SSR) and six genomic (G)-SSR markers revealed monomorphic bands in micropropagated shoots and plants in lingonberry hybrid H1 and cultivar Erntedank conforming genetic integrity. Epigenetic variation was studied by quantifying cytosine methylation using a methylation-sensitive amplification polymorphism (MSAP) technique. DNA methylation ranged from 32% in greenhouse-grown hybrid H1 to 44% in cultivar Erntedank under a tissue culture system. Although total methylation was higher in in vitro grown shoots, fully methylated bands were observed more in the greenhouse-grown plants. On the contrary, hemimethylated DNA bands were more prominent in tissue culture conditions as compared to the greenhouse-grown plants. The study conclude that lingonberry maintains its genetic integrity but undergoes variable epigenetic changes during in vitro and ex vitro conditions.

Opportunities and Challenges of In Vitro Tissue Culture Systems in the Era of Crop Genome Editing.

Currently, the development of genome editing (GE) tools has provided a wide platform for targeted modification of plant genomes. However, the lack of versatile DNA delivery systems for a large variety of crop species has been the main bottleneck for improving crops with beneficial traits. Currently, the generation of plants with heritable mutations induced by GE tools mostly goes through tissue culture. Unfortunately, current tissue culture systems restrict successful results to only a limited number of plant species and genotypes. In order to release the full potential of the GE tools, procedures need to be species and genotype independent. This review provides an in-depth summary and insights into the various in vitro tissue culture systems used for GE in the economically important crops barley, wheat, rice, sorghum, soybean, maize, potatoes, cassava, and millet and uncovers new opportunities and challenges of already-established tissue culture platforms for GE in the crops.

Development of tissue culture system with automated pulsation and Kalman filter control for an artificial artery model.

Tissue-engineered arterial vessels have been used as substitutes for unnecessary animal experiments to evaluate the pharmacokinetics of drugs targeting various arteriopathies caused by structural or physiological arterial defects. An arterial tissue culture system was established to simulate the mechanical characteristics of a heart-beating pump and to do online feedback control of lactate and glucose concentrations. The mechanically controlled flow pump mimicked the heart pumping inside a tissue-engineered artery composed of muscle and endothelial cells within a nanofibrous scaffold. After monitoring the pH of the culture medium online, lactate and glucose were estimated using the Kalman filter algorithm, and the set-point online control was operated to maintain glucose for artery tissue engineering. The composition of the artificial artery was confirmed by immunofluorescence staining, and its mechanical characteristics were examined. The online automated system successfully demonstrated its applicability as a standardized process for arterial tissue culture to replace animal arterial experiments.

In vitro regeneration of Haloxylon ammodendron

Haloxylon ammodendron (C.A.Mey) Bunge is one of the important species of arid desert vegetation in China, and it is also an ideal material for studying plant stress resistance, which plays an extremely important role in protecting desert ecosystems and preventing land desertification. However, there are relatively few studies on the regeneration of the fusiform, and the genetic transformation system has not been successfully reported, which restricts the in-depth study of the molecular mechanism of shuttle propagation and stress resistance. In the present study, the seeds, hypocotyls, cotyledons, cotyledon nodes, terminal buds and fixed buds were used as explants, and a set of tissue culture and plant regeneration system was established by inducing adventitious buds, adventitious bud rooting and transplanting. The results showed that amongst different H. ammodendron explants, i.e., seeds, hypocotyls, cotyledons and cotyledon nodes, the last induced budding effect was better. The optimal medium for inducing clandine buds by cotyledon segment differentiation is 0.5 mg·L-1 NAA+0.5 mg·L-1 6-BA induced budding rate was high, reaching 61.90%, rooting medium was 1/2 MS+1 mg·L-1 NAA +1 mg·L-1 IBA + 1 mg·L-1 IAA with a rooting rate of 50%. The results of this study will provide a theoretical basis for the genetic transformation of H. ammodendron.