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Plant Tissue Culture Media
Abstract
Some plants are able to synthesize the essential requirements of vitamins for their growth. Some vitamins are required for normal growth and development of plants, they are required by plants as catalysts in various metabolic processes. They may act as limiting factors for cell growth and differentiation when plant cells and tissues are grown Invitro [2]. The vitamins most used in the cell and tissue culture media include: thiamin (B1), nicotinic acid and pyridoxine (B6). Thiamin is necessarily required by all cells for growth [11]. Thiamin is used at concentrations ranging from 0.1 to 10 mg.l⁻¹. Nicotinic acid and pyridoxine, however not essential for cell growth of many species, they are often added to culture media [12]. Nicotinic acid is used at a concentration range 0.1-5 mg.l⁻¹ and pyridoxine is used at 0.1-10 mg.l⁻¹. Other vitamins such as biotin, folic acid, ascorbic acid, pantothenic acid, tocopherol (vitamin E), riboflavin, p-amino-benzoic acid are used in some cell culture media however, they are not growth limiting factors. It was recommended that vitamins should be added to culture media only when the concentration of thiamin is below the desired level or when the cells are required to be grown at low population densities [14]. Although it is not a vitamin but a carbohydrate, myo-inositol is added in small quantities to stimulate cell growth of most plant species [13]. Myo-inositol is believed to play a role in cell division because of its breakdown to ascorbic acid and pectin and incorporation into phosphoinositides and phosphatidyl-inositol. It is generally used in plant cell and tissue culture media at concentrations of 50-5000 mg.l⁻¹.
... Culture media generally consist of inorganic elements and organic compounds. The composition and types of media used for the HR culture of plants influence SMs production (George et al., 2008;Saad & Elshahed, 2012). In previous studies, it was proclaimed that the selection of the suitable basal media for HR induction and growth for the accumulation of desirable products is mainly based on the ideal medium and plant species (Washida et al., 1998;Dhakulkar et al., 2005;Kumar et al., 2006). ...
... The accumulation of SM is highly influenced by several factors like medium salt concentration, the type and quantity of carbohydrates, phosphate, and nitrate, and the growth regulator levels (Washida et al., 1998;Dhakulkar et al., 2005;Kumar et al., 2006;George et al., 2008;Kim et al., 2012;Saad & Elshahed, 2012). It has been reported that different formulations of media such as B5, Linsmaier and Skoog (LS), MS, and SH, have been used for different plant cultures (Murthy et al., 2014). ...
Brassica juncea is a vegatable that are rich in glucosinolate (GSL) content. The hairy root (HR) cultures system is one of the most useful tools for secondary metabolites (SM) biosynthesis under various growth conditions. In the past, GSLs were mostly used as biopesticides in agriculture, anti-nutritional factors in fodder, and flavors in condiments. However, in recent days, GLSs have received much attention in human health. To investigate the growth response and variation of GSLs accumulation, HRs of mustard were grown in different growth media and auxins. The HRs growth pattern varied largely under the treatments of growth media and auxin. The full-strength SH media responded greatly for achieving the highest dry weight (DW) followed by the ½ SH media and the lowest DW was obtained in full-strength MS media. In all the auxin treatments the HRs production was higher than that of the control. It was noted that at higher NAA and IBA concentrations HR production was increased than that at the lower concentrations. In addition, different growth mediums significantly influenced the GSLs accumulation in mustard HR. The results revealed that ½ B5 media showed the highest total GSLs content followed by B5 and ½ SH. Treatment of mustard HRs with auxins such as IAA and IBA negatively influenced the accumulation of GSLs except for 4-methoxyglucobrassicin. We, therefore, suggest that HRs are a viable option for improving the GSLs content from the HR culture of mustard and that SH and ½ B5 medium provides an alternative approach for mass production of HRs and GSLs in mustard, respectively.
... The success rate of plant tissue culture depends on the choice of the nutrient medium. In fact, the cells of most plant cells can be grown in cultured media (Saad and Elshahed, 2012). Plant growth regulators are the basic media parts in deciding the formative pathway of plant cells. ...
... Optimal growth and morphogenesis of tissues may vary for different plants according to their nutritional requirements. Additionally, tissues from diverse parts of plants may have different requirements for satisfactory growth (Saad and Elshahed, 2012). The study of MD2 variety pineapple has been done by Danso et al. (2008) through direct in vitro micropropagation which displayed optimum result for in vitro proliferation when the concentration of NAA in the medium was 2.0 mg/L and the BAP 5.0 mg/L. ...
Pineapples (Ananas comosus L. Merr) are fruits that belong to the Bromeliaceae family. Pineapple variety MD2 is one of the varieties that has gained a place in the market among pineapple farmers due to its high value and quality. However, it is difficult to meet the demand for planting materials using conventional propagation techniques. Hence, plant tissue culture technology is one of the methods that has been widely used in the agriculture industry that boosts up the production of pineapple planting materials within a short period and is cost-efficient. The objective of this study was to determine the effect of plant growth regulator concentration to in vitro culture of MD2 variety pineapple. In this study, the various concentrations of 6-Benzylaminopurine (BAP) and α-Naphthalene Acetic Acid (NAA) for in vitro culture of MD2 pineapple were studied. The plantlets were effectively initiated from MD2 pineapple crown on Murashige and Skoog (MS) basal salt containing 30.0 g/L sucrose, and 2.0 mg/L BAP in two months of culture. Next, the pineapple plantlet was subculture on shooting medium containing full strength solid Murashige and Skoog (MS) medium supplemented with vitamins with various concentration BAP (0, 1.0, 2.0, 3.0, 4.0, 5.0, 6.0, 7.0, and 8.0 mg/L) and NAA (0, 1.0, and 2.0 mg/L). The result obtained showed that the solid MS medium added with 30.0 g/L sucrose, without any BAP and NAA (T1) had the highest in vitro shoot regeneration. Meanwhile, the solid MS medium with 30.0 g/L sucrose with 1.0 mg/L NAA (T1) recorded the highest plantlet height (cm). The mean value for in vitro shoot regeneration in T1 and plantlet height (cm) in T1 were 2.80 (±0.5) and 4.40 (±0.3). To conclude, less amount plant hormone regulator required to obtain the mass quantity of in vitro clonal pineapple that can help solve the problem of lack of plant material in the pineapple crop industry.
... The gelling agent most used in-vitro studies is agarose, a polysacharide generally extracted from seaweed. According to Saad and Elshahed (2012), agarose contains a low percentage Ca, Mg, K, and Na. ...
... Sugar or sucrose is used as an alternative energy source applied in the culture medium. Sucrose is the best energy-producing carbohydrate than other sources like lactose, maltose, galactose, mannitol and technical sugar (Saad and Elshahed 2012;Placide et al. 2012). A side from being a source of energy, sugar also serves as the osmotic pressure of the media. ...
Fitriani H, Aryaningrum PD, Hartati NS. 2016. Proliferation of embryogenic callus of Satoimo taro (Colocasia esculenta var. antiquorum) in culture media with various level of sucrose and gelling agent. Nusantara Bioscience 8: 316-320. Satoimo taro (Colocasia esculenta var. antiquorum) is one of the promising varieties of taro which have potentially high value to cultivate. Tissue culture or in-vitro culture considers an economically useful technique to mass propagates this variety of taro. The research was conducted to figuring out the optimum media for proliferation success of embryogenic callus of Satoimo. The explants used embryogenic callus of 12-months-old storage materials obtained from the previous study. Proliferation media include a basal half-strength MS salt (Murashige and Skoog) enriched with of 0.1 mg/L TDZ, 0.05 mg/L 2.4 D, and 100 mg/L L-glutamine. Adding to the basal media, we set the treatments by combining two factors i.e. Sucrose with three levels of concentration i.e. 3, 4, and 5% which cross-tabulated with three gelling agents, i.e., Agarose, Phytagel, and Gerlite. The research parameters included the percentage of callus diameter growth, the color and callus discoloration, and the structure of callus. The data recorded periodically every 7 d in one month. Data were analyzed using analysis of variance and followed by Duncan Multiple Range Test (DMRT) to determine the significant difference (p<0.05) among the treatments. The result revealed that the callus diameter growth has the greatest percentage at medium with a combination of Phytagel and sucrose 4%. The Micro-agar enhanced the presence of callus with yellow color. Phytagel combined with a high percentage of sucrose (5%) indicated browning properties of callus. Gelrite causes the callus with discoloration appearance (white).
... Shikonin, a red naphthoquinone derivative, is a secondary metabolite that specifically occurs in boraginaceous plants, and is the active principle of the medicinal plant Lithospermum erythrorhizon (17). Since this compound and its derivatives exhibit antibacterial activity, their functions as phytoalexins have also been reported (18). ...
... Since this compound and its derivatives exhibit antibacterial activity, their functions as phytoalexins have also been reported (18). The biosynthesis of shikonin includes a key prenylation step catalyzed by geranyl diphosphate (GPP) 1 :4-hydroxybenzoate (4HB) 3-geranyltransferase; i.e. coupling of the shikimate and mevalonate pathways (17,19,20). This enzyme plays a critical role in the regulation of shikonin biosynthesis in cell cultures of L. erythrorhizon, i.e. up-and down-regulation of this enzyme activity directly affects the production of shikonin (21,22). ...
Two cDNAs encoding geranyl diphosphate:4-hy- droxybenzoate 3-geranyltransferase were isolated from Lithospermum erythrorhizon by nested PCR using the conserved amino acid sequences among polyprenyl- transferases for ubiquinone biosynthesis. They were functionally expressed in yeast COQ2 disruptant and showed a strict substrate specificity for geranyl diphosphate as the prenyl donor, in contrast to ubiquinone biosynthetic enzymes, suggesting that they are involved in the biosynthesis of shikonin, a naphthoquinone secondary metabolite. Regulation of their expression by various culture conditions coincided with that of geranyltransferase activity and the secondary metabolites biosynthesized via this enzyme. This is the first established plant prenyltransferase that transfers the prenyl chain to an aromatic substrate.
... The required type, concentration, and time of disinfectant exposure differ for different plants and plant parts (Dar et al., 2012), and the optimal percentage of NaOCl for sterilization must be determined. Most tissue culture studies found that low levels of auxins induced rooting, while high levels induced callus formation (Saad and Elshahed, 2012). At the cellular level, the cell division cycle is controlled by the joint action and ratio to each other of auxins and cytokinins (George et al., 2008). ...
... The morphogenesis and growth of plant tissue under in vitro conditions depend on the composition of the nutrient medium: macro-and micro-salts, vitamins, carbohydrates, amino acids, and plant growth regulators (PGRs) [24]. Amino acids provide an essential quick nitrogen source in plants as compared with inorganic nitrogen [25]. The amino acid asparagine produced the healthiest vigorous shoots, the lowest withering percentages, and took a maximum number of days to show withering of Rosa centifolia shoots [26]. ...
The use of nanomaterials in biotechnology for the in vitro propagation of medical plants and the accumulation of certain biologically active metabolites is becoming an efficient strategy. This study aimed to evaluate the influence of the concentration (0, 1, 10, 50, and 100 mg L−1) of two types of nanofibers on the growth characteristics, the antioxidant status, and the production of steviol glycosides in micropropagated Stevia rebaudiana Bert. plantlets. The nanofibers were synthesized by aspartic acid derivatives (L-Asp) Ag salts self-organized into nanofibers with two different molecular structures: monomeric, containing one residue of L-Asp with one hydrophilic head which bonds one Ag ion (NF1-Ag salt); and dimeric, containing two residues of L-Asp with two hydrophilic heads which bond two Ag ions (NF2-Ag salt). An increase in the shoots from the explants’ number and length, biomass accumulation, and micropropagation rate was achieved in the plants treated with the NF1-Ag salt in concentrations from 1 to 50 mg L−1 after 30 days of in vitro proliferation compared to the NF2-Ag salt. In contrast, the plants grown on MS media supplemented with NF2-Ag salt exhibited an increase in the level of stevioside, rebaudioside A, and mono- (CQA) and dicaffeoylquinic (DCQA) acids as compared to the NF1-Ag salt.
... Plant tissue culture media should have all the essential nutrients like minerals, carbon supplements, vitamins, and other organic supplements, PGRs (plant growth regulators), solidifying-agents, and many more for the proper growth of the entire plant or production of any desired secondary metabolite from plants (Saad and Elshahed 2012). Basal media, those are often employed in tissue culture techniques, are MS (Murashige and Skoog) medium, B 5 (Gamborg) medium, LS (Linsmaier and Skoog) medium, and NN (Nitsch and Nistch) medium (Murashige and Skoog 1962;Linsmaier and Skoog 1965;Gamborg et al. 1968;Nitsch and Nistch 1969). ...
Genus Crinum L. is a member of the Amaryllidaceae family having beautiful, huge, ornamental plants with umbels of lily-like blooms that are found in tropical and subtropical climates all over the world. For thousands of years, Crinum has been used as a traditional medicine to treat illnesses and disorders. Numerous distinct alkaloids of the Amaryllidaceae group, whose most well-known properties include analgesic, anticholinergic, antitumor, and antiviral, have recently been discovered by phytochemical analyses. However, because of decades of overexploitation for their economically significant bioactive ingredients and poor seed viability and germination rates, these plants are now threatened in their native environments. Because of these factors, researchers are investigating micropropagation techniques to optimize phytochemicals in vitro. This review’s objective is to offer details on the distribution, phytochemistry, micropropagation, in vitro galanthamine synthesis, and pharmacology which will help to design biotechnological techniques for the preservation, widespread multiplication, and required secondary metabolite production from Crinum spp.
Key points
• Botanical description and phytochemical profile of Crinum spp.
• In vitro micropropagation method of Crinum sp.
• Bioactive compound galanthamine isolation techniques and its pharmacological properties.
... In addition, organic nitrogen sources in MS media may play an essential role in the multiplication of sugarcane shoot apical meristem. Some amino acids such as asparagine, cysteine, casein, glutamine, and glycine are primarily used in culture media as organic nitrogen sources (Saad and Elshahed 2012). The addition of 100 ppm glutamine and 2 ppm glycine into MS media produced robust and healthy sugarcane plantlets (Sugiharto, unpublished data). ...
Sugarcane is grown by small, medium, and large rural farmers in several countries around the globe. In addition, the primary objective is to increase cane yield, sugar recovery and sustainably improve the livelihoods of cane farmers. For producing a large amount of biomass and sugarcane, crop extracts a large amount of nutrients from the soil and accumulate in the plant. The regular harvesting of natural resources consequently from the soil mitigates a high amount of nutrients. Therefore, there is always a need to replace these nutrients with other sources of fertilization. Soil textural properties and fertility status under changing climatic conditions also play an important role. Several alternatives can be utilized to increase the sustainable nutrient use efficiency of both macro and micronutrients to make a balance for the profitability of the crop. Two of these natural alternatives are the use of green and organic manure, i.e., press mud and farmyard manure. This chapter aims to develop an integrated nutrient management approach for the global cane farmers that would improve the quality and productivity of the canes and improve water, nutrients, and pesticide use efficiencies.
... In addition, organic nitrogen sources in MS media may play an essential role in the multiplication of sugarcane shoot apical meristem. Some amino acids such as asparagine, cysteine, casein, glutamine, and glycine are primarily used in culture media as organic nitrogen sources (Saad and Elshahed 2012). The addition of 100 ppm glutamine and 2 ppm glycine into MS media produced robust and healthy sugarcane plantlets (Sugiharto, unpublished data). ...
Sugarcane is considered as an important industrial crop to produce sugar, and nearly 80% of sugar production worldwide is produced from this plant. Sugarcane is a C4 plant that has a higher photosynthetic potential. Abiotic and biotic stresses have a diverse impact on the growth and productivity of sugarcane. Understanding the biochemical and physiological mechanism of these stresses is one of the most important aspects to improve the variety of plants that can meet better quality and quantum. Progress in the development of new sugarcane cultivars by conventional breeding has been hindered by its complex polyploid-aneuploid genome leading to a long breeding period. These types of constraints offer an opportunity to generate new sugarcane cultivars through biotechnological approaches. The new variety of sugarcane with desirable traits, such as drought tolerant and virus resistance, have been attempted to increase the yield of the plant. The inducing accumulation of compatible solutes such as sugar and betaine help sugarcane to adapt and survive in water limited environment. Biotic stress causes a significant loss in sugarcane growth and yield. Pathogen-derived resistance (PDR) and RNA interference (RNAi) technologies have been applied to engineered sugarcane cultivars having resistance to the sugarcane mosaic virus. In addition, genetic engineering of sucrose metabolism is also an important means to control carbon flux through the enzyme sucrose-phosphate synthase, which is responsible for the synthesis of sucrose. Here, we summarize recent developments in the biotechnological approaches to improve sugarcane yield by developing stress tolerance efficiency, increased yield, and virus resistance, including potential and challenges of genome editing technological applications.
... Unlike inorganic nitrogen, amino acids are easily assimilated by plant cells and tissues [50]. Saad and Elshahed [51] suggested that plant cells have a higher capacity to take up and transport nitrogen from organic sources rather than inorganic ones. Several studies have reported using amino acids as an organic nitrogen source during in vitro propagation of several species such as Fragaria × Ananassa duch cv. ...
The growing demand for native planting material in ecological restoration and rehabilitation for agro-silvo-pastoral ecosystems has resulted in a major global industry in their sourcing, multiplication, and sale. Plant tissue culture is used for producing high-quality, disease-free, and true-to-type plants at a fast rate. Micropropagation can help to meet the increasing demand for planting material and afforestation programs. However, in vitro plant propagation is an expensive technique compared to conventional methods using suckers, seeds, and cuttings. Therefore, adopting measures to lower production costs without compromising plant quality is essential. This can be achieved by improving the culture media composition. Incorporating organic growth additives can stimulate tissue growth and increase the number of shoots, leaves, and roots in culture media. Organic growth supplementation speeds up the formation and development of cultures and yields vigorous plants. Plant regeneration from meristems (shoot tips and axillary buds) is a reliable way to produce true-to-type plants compared with callus and somatic embryogenesis regeneration, but in vitro culture environments can be mutagenic. Therefore, detecting somaclonal variations at an early stage of development is considered crucial in propagating plants. The genetic stability of in vitro regenerated plants needs to be ascertained by using DNA-based molecular markers. This review aims to provide up-to-date research progress on incorporating organic growth additives to enhance in vitro tissue culture protocols and to emphasize the importance of using PCR-based molecular markers such as RAPD, ISSR, SSR, and SCoT. The review was assessed based on the peer-reviewed works published in scientific databases including Science Direct, Scopus, Springer, JSTOR, onlinelibrary, and Google Scholar.
... In addition, organic nitrogen sources in MS media may play an essential role in the multiplication of sugarcane shoot apical meristem. Some amino acids such as asparagine, cysteine, casein, glutamine, and glycine are primarily used in culture media as organic nitrogen sources (Saad and Elshahed 2012). The addition of 100 ppm glutamine and 2 ppm glycine into MS media produced robust and healthy sugarcane plantlets (Sugiharto, unpublished data). ...
This edited volume focuses on the core aspects of sugarcane production-management under stressful environments as well as innovative strategies for augmenting crop growth & productivity through intrinsic and extrinsic manipulations. The various chapters aim at bringing out comprehensive and advance information on different aspects of sugarcane cultivation under stress environments and impact of climate change on the sustainability of sugarcane production. The book encompasses information about crop production management, physiological & nutritional requirements, ratooning, ripening and post-harvest losses management. It also delineates various technologies that support the continued use and improvement of sugarcane as renewable source of food, fiber and bio-energy. The manipulations at cellular and molecular levels, molecular breeding approaches and post-harvest technologies are also included. The area under sugarcane cultivation is gradually increasing because of its diversification potential. The high productivity and biomass of the cane crop also makes it a key source for use as bio-energy crop and a promising raw material for bio-based agro-industries. However, poor crop & biomass productivity due to abiotic stress is the foremost constraint in its future commercial exploitation as sustainable feed-stock for bio-based industries. It is therefore imperative to understand the cellular-molecular modulation responsible to productivity barrier under specific stress situation(s) for better sugarcane quality and quantum under field condition. Some of these innovative approaches are delineated in this book. This book is of interest to progressive sugarcane growers, millers, industrial entrepreneurs, sugarcane scientists, cane development and extension officers, sugar industry managers and valuable source of reference worldwide.
... Besides carbon source, plant growth regulator (PGR) is also an important element in plant tissue culture since it plays vital roles in the stem elongation, tropism, and apical dominance. In tissue culture, auxins are usually used to stimulate callus production and cell growth, initiate shoots and rooting, induce somatic embryogenesis and stimulate growth from shoot apices and shoot stem culture (Saad & Elshahed, 2012). Initial inoculum density is a key factor in establishing cell suspension cultures of any plant species because cells cannot resume and even stop their active growth after the transferring and subculturing processes if the initial inoculum is below the critical cell density. ...
Cell suspension culture of Citrus suhuiensis (C. suhuiensis) was established to determine the best growth condition based on the effects of carbon source, plant growth regulators (PGRs) and inoculum size. Friable callus from the cotyledon explant was used to initiate the cell suspension cultures. Murashige and Skoog (MS) medium supplemented with 50 g/L glucose achieved the highest cell dry weight (CDW) and µmax of 23.2 g/L and 0.47 h-1 respectively compared to glucose at 30 g/L and other carbon sources. For PGRs effects, 0.5 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D) gave the highest amount of CDW and µmax; 11.91 g/L and 0.32 h-1. Furthermore, the inoculum size at 10% (v/v) gave the maximum CDW and µmax of 15.45 g/L and 0.71 h-1 respectively compared to 20% (v/v) and 30% (v/v) of inoculum. The results achieved can be used for further statistical optimization of C. suhuiensis cell suspension culture growth parameters in which the interactions between those parameters can be further studied and understood.
... Myo is also commonly added in small quantities to the culture media in order to stimulate cell growth. The metabolic route of this vitamin leads to its breakdown to ascorbic acid and pectin (Saad and Elshahed, 2012). For this reason, Myo is considered, together with Thia, an essential organic compound for plant tissue cultures (George et al., 2008). ...
Mineral nutrients are essential components of basal media for optimal growth and development of plants grown in vitro. Their deficiency causes physiological disorders. Vitamins have several physiological functions, however, their role in the growth and development of micropropagated plants remains scarce and controversial. The objective of this work has been to get insight on the role of mineral nutrients and vitamins as main components of the in vitro basal medium on the quality of micropropagated kiwiberry (hardy kiwi). Two computer tools were used: design of experiments (DOE) and machine learning (ML), particularly neurofuzzy logic (NL). Two different experiments were designed using a well-sampled five-dimensional experimental design space of 33 treatments using DOE. In both experiments five independent factors were established, in the first the following mineral salts: NH4NO3, KNO3, Mesos, Micros and iron, while in the second, the following five vitamins: myoinositol, thiamine, nicotinic acid, pyridoxine and vitamin E. Neurofuzzy logic models were used to identify the critical vitamins and mineral nutrients on shoot length (SL) and shoot quality (SQ). Neurofuzzy logic models showed that only 7 out of 18 mineral elements were critical factors affecting both, SL and SQ, while only 3 out of 5 vitamins were relevant. In conclusion, these results shed light on the effect of mineral nutrients as key requirements on shoot growth and quality and, for the first time, identified the relevant role of vitamin E on shoot growth of micropropagated kiwiberry.
... In this study, Murashige & Skoog (MS) basic media was used with good growth results in all three types of Begonia explants. MS media contains nitrate, ammonium, calcium, and other macro and microelements that can affect the growth of explants (Saad & Elshahed, 2012). MS media was combined with cytokinin group PGR, namely BA and TDZ. ...
Giam Siak Kecil Bukit Batu is a biosphere reserve which one of its functions is as a habitat for wildlife. However, biodiversity data in the Giam Siak Kecil Bukit Batu Biosphere Reserve (GSKBB-BR) is still very minimal, including insects (Coleoptera and Hemiptera). This research was conducted to determine the diversity of Coleoptera and Hemiptera in the GSKBB Biosphere Reserve, Riau, Indonesia. The research was carried out using an exploratory method using "lights trap". The results of the study found 30 species, from 11 families of the order Coleoptera (23 species) and Hemiptera (7 species) in the GSKBB-BR. The diversity index of Coleoptera and Hemiptera at the observation site was moderate (H = 2.73), with a high evenness index (0.80). Scarabaeidae (order Coleoptera) is the family with the highest number of species found (8 species), while the most abundant species were Tibicen linnei and Pomponia fusca (Cicadidae/Hemiptera). Based on their functional roles, Coleoptera and Hemiptera with the highest number are herbivores (17 species), followed by predators (7 species) and decomposers (3 species). The range of values for temperature and humidity at the research site are in normal conditions. The GSKBB-BR area is an important remaining habitat for wildlife in Riau, including various types of insects (Coleoptera and Hemiptera); whose potential still needs to be revealed, and must be managed properly.
... In this study, Murashige & Skoog (MS) basic media was used with good growth results in all three types of Begonia explants. MS media contains nitrate, ammonium, calcium, and other macro and microelements that can affect the growth of explants (Saad & Elshahed, 2012). MS media was combined with cytokinin group PGR, namely BA and TDZ. ...
Three species of Begonias endemic to Java and Sumatra, namely Begonia leuserensis, Begonia atricha and Begonia scottii, were conserved in Cibodas Botanic Gardens as sources of germplasm for ornamental plant and/or medicines. However, the information on efficient hormones concentration and their culture media application through an in vitro propagation effort is still limited. Therefore, this study aimed to explain the growth response of three species of Begonias using various hormones concentrations and culture media through in vitro propagation. The culture media using Murashige & Skoog (MS) media that combinedwith 6-Benzyladenine (BA) dan Thidiazuron (TDZ) hormones in different concentrations i.e. 0.5 mg/L, 1 mg/L, 2 mg/L, and 3 mg/L. Observation parameter included shoot number, plantlets height, and leaves number. The data were analyzed using analysis of variance (ANOVA) with the F test at a 5% significance level. The results showed that three species of Begonias were observed to have different growth responses in the combination of MS+BA and MS+TDZ media. The combination of MS+TDZ media produces more shoots number, while the combination of MS+BA media influenced higher in leaves number. A concentration of 0.5 mg/L of hormone showed a good regeneration, therefore were recommended for in vitro propagation of Begonia species.
... For shoot induction, BAP (from the class of cytokinin) with different concentrations was tested. Cytokinin is a common plant growth regulator for shoot induction, cell division and cell proliferation in tissue culture process (Saad and Elshahed, 2012). The usage of BAP as plant growth regulator for F. deltoidea shoot induction has been supported by other researchers, which used BAP singly or in combination with NAA for in vitro shoot regeneration of F. deltoidea nodal explant (Abdullah et al., 2015). ...
Ficus deltoidea or commonly known as ‘mas cotek’ is a herbal plant indigenous to Southeast Asia including Malaysia and Indonesia. This plant is popular for its medicinal values such as improve blood circulation, regain energy and enhance fertility naturally for both men and women. The main objective of this study is to develop in vitro clonal propagation method for rapid production of F. deltoidea using different concentrations of benzyl aminopurine (BAP) through shoot induction and multiplication, rooting and subsequent establishment in soil following acclimatization. Surface sterilization of the leaf explants was done using mercury chloride and ethanol as the disinfectants. Pre-treatment of the explants with carbendazim successfully reduced the occurrence of fungal contamination. At the end of the experiment, no shoot and root induction were observed but calli were successfully induced on MS medium containing 1.0, 1.5, 2.0, 2.5 and 3.0 mg/l BAP, with calli induced from 3.0 mg/l BAP were bigger and healthier. In short, the higher the concentration of BAP used, the higher tendency for the explant to induce callus.
... Similar results were reported in germination of seeds of L. pumila due to the presence of BAP in half-strength MS medium (Hartinie and Jualang, 2007). BAP or cytokinin promotes cell division and regulates growth and development in plant tissue under certain condition (Saad and Elshahed, 2012). It is also more commonly used than kinetin for inducing rapid multiplication of shoots, buds, or meristems (Chawla, 2002). ...
Labisia pumila or commonly known as Kacip Fatimah is one of the plants that were used as a medicinal purpose. Previously, several studies on the regeneration of the plant through tissue culture had been conducted. L. pumila can be planted using macropropagation technique in which L. pumila can be propagated by using stem cuttings. However, the seed yields were low. This study was initiated to regenerate L. pumila using micropropagation from the embryo as the explant. Embryos were cultured on different strength MS culture mediums which are full–strength and half–strength whereby the half-strength MS medium was supplemented with 0.5 ppm benzylaminopurine (BAP) for shoot induction. Shoot formation was achieved from both media. High rate of shoot formation occurred on full-strength MS medium without plant growth regulators. Multiple shoots was established on half-strength MS medium containing 0.5 ppm BAP. Shoots elongation and plantlet establishment were produced with culture on half-strength MS medium with combination of 0.5 ppm BA and 0.5 ppm IAA.
... Sowing of orchid seeds in the laboratory requires nutrients that are packaged in a basic medium. It is known that there are several basic media commonly used, namely MS media (Murashige and Skoog) for micropropagation of orchids in general, NP (New Phalaenopsis) for micropropagation of the Phalaenopsis orchids genera, and VW (Vacin and Went) for micropropagation of orchids from Dendrobium genera [22]. In vitro propagation requires a growing medium that contains both macro and micro-nutrients that support plant growth and aseptic conditions [23]. ...
Vanda tricolor Lindey var. Suavis is an endemic orchid from Mt.Merapi, Yogyakarta Special Region, Indonesia. This orchid has beautiful flowers with unique patterns of white and purple spots, fragrant and can live in the slopes of Mt.Merapi which is a very active volcano. UGM in collaboration with the Netherlands carried out ex situ conservation of the V. tricolor Merapi through the self-pollination of orchids by the Queen of the Netherlands, Queen Maxima on March 11, 2020 at the UGM campus, Yogyakarta. In honor, the new generation of crosses is named Vanda tricolor var Suavis ‘Queen Maxima’. This study aims to produce mass quantities of the orchid and characterize the HSP70 gene in it. Methods: Standard in vitro culture for Vanda on MS, NP and VW medium, isolation of plant gDNA, PCR with V. tricolor HSP70primers and sequencing of amplicon DNA. The results showed that > 90 % of V. tricolor ‘Queen Maxima’ seeds germinated well in all media and the best on VW medium. V. tricolor has the HSP70 protein with PTZ00009 super family amino acid motif that 87 % similar to the HSP70 protein from the Phalaenopsis equestris orchid, which is probably the reason V. tricolor become superior to high temperature environments.
... In Sri Lanka Tall coconuts, 6% (w/v) sucrose was used in MS medium for 15 to 17 d (Fernando et al. 2009). Sucrose is expected to result in increased plumule growth since its pure form induces near optimal rates of growth as a widespread transport molecule (Mello et al. 2001;Placide et al. 2012;Saad and Elshahed 2012). Sucrose has high solubility in water, electrically neutral, and is shown to have no inhibitory effects on majority of biochemical processes (Placide et al. 2012). ...
... As a rule, culture media components contain, as average, 18 different mineral nutrients, some required at high concentrations (macronutrients) such as nitrogen, potassium, calcium, phosphorus, sulfur and magnesium, while others are required at lower concentrations (micronutrients), such as manganese, zinc, boron, molybdenum, copper and iron, among others, being all essential for certain physiological processes (Twaij et al., 2020). Together with mineral nutrients, a source of carbon, normally sucrose, as well as other organic molecules, such as vitamins and amino acids, some plant growth regulators, are supplied to media to ensure a healthy plant growth and development (Saad and Elshahed, 2012). In addition, there are additional factors that show a significant impact on mineral nutrition, such as the genotype, because even closely related species have been shown to present differential behaviors toward certain media ingredients Nezami-Alanagh et al., 2014). ...
Plant nutrition is a crucial factor that is usually underestimated when designing plant in vitro culture protocols of unexploited plants. As a complex multifactorial process, the study of nutritional imbalances requires the use of time-consuming experimental designs and appropriate statistical and multiple regression analysis for the determination of critical parameters, whose results may be difficult to interpret when the number of variables is large. The use of machine learning (ML) supposes a cutting-edge approach to investigate multifactorial processes, with the aim of detecting non-linear relationships and critical factors affecting a determined response and their concealed interactions. Thus, in this work we applied artificial neural networks coupled to fuzzy logic, known as neurofuzzy logic, to determine the critical factors affecting the mineral nutrition of medicinal plants belonging to Bryophyllum subgenus cultured in vitro. The application of neurofuzzy logic algorithms facilitate the interpretation of the results, as the technology is able to generate useful and understandable “IF-THEN” rules, that provide information about the factor(s) involved in a certain response. In this sense, ammonium, sulfate, molybdenum, copper and sodium were the most important nutrients that explain the variation in the in vitro culture establishment of the medicinal plants in a species-dependent manner. Thus, our results indicate that Bryophyllum spp. display a fine-tuning regulation of mineral nutrition, that was reported for the first time under in vitro conditions. Overall, neurofuzzy model was able to predict and identify masked interactions among such factors, providing a source of knowledge (helpful information) from the experimental data (non-informative per se), in order to make the exploitation and valorization of medicinal plants with high phytochemical potential easier.
... Growth regulator substances which are often used in in vitro culture are auxin and cytokinin. Auxins, in tissue culture, are usually used to stimulate callus production and cell growth, to initiate shoots and rooting, to induce somatic embryogenesis, to stimulate growth from shoot apices and shoot stem culture [11]. Auxins play roles in directly affect processes of stimulating cell elongation and cell division, whereas cytokinin involved in cell growth and differentiation, apical dominance, axillary bud growth and leaf senescence. ...
Paraphalaenopsis is a small genus of the tribe Vandeae, the genus comprises 4 species: Paraphalaenopsis denevei, Paraphalaenopsis laycockii, Paraphalaenopsis labukensis and Paraphalaenopsis serpentilingua. All are known endemic to Borneo, and also rare due to over exploitation, therefore requires rescue efforts. In vitro culture is one of effort that can be done to regenerate a large number of seedlings and to ex situ conservation. Optimal growth and morphogenesis of tissues may vary for different plants according to their nutritional requirement. Auxin (NAA) is an essential factor for root growth in tissue culture work. Activated charcoal is often used in tissue culture to improve cell growth and development. This research was conducted to find out the in vitro growth effect from leaf explants of P. labukensis with plant growth regulators (NAA) and activated charcoal on ½ MS medium as basal medium. The treatment with NAA 2 ppm and 1 g/l activated charcoal showed the highest percentage of life (50 %) and percentage of leaf (57.1 %). The largest percentage of roots showed in 2 g/l activated charcoal (35.7 %). The death of leaf explants P. labukensis was caused by browning. The phenolic compound which accumulated during wounding is causing browning.
... The success of in vitro technique as a tool for plant propagation much depends upon the characteristics of medium used (Saad and Elshahed, 2012;Arab et al., 2014). In principle, basic requirement of explants cultured in vitro is similar to intact plants. ...
This study was aimed at investigating the effect of NAA and BAP on the development of basal slip slices in tissue culture of pineapple cv. Tangkit. The experiment was conducted at the Plant Biotechnology Laboratory Agricultural Faculty University of Jambi from July through to October 2016. Six level of NAA concentrations (0, 1, 2, 3 , 4 and 5 mgL-1) were tested in combination with six levels of BAP concentration (0, 1, 2, 3 , 4 and 5 mgL-1). A Completely Randomized Design with 5 replicates was employed in this trial. Data on the percentage of explants forming shoots, time required for shoot formation, number of shoots growing per explants, and callus proliferation were recorded. Results showed that without the involvement of plant growth regulators there had been no growth on cultured explants. However, shoot growth was found on explants cultured on medium supplemented with 1 – 3 mgL-1 NAA or 1 – 5 mgL-1 BAP. The application of NAA higher than 3 mgL-1 was found to suppress shoot growth even in combination with BAP. Shoot formation was found to be fast on medium supplemented with 1 mgL-1 NAA + 4 mgL-1 BAP, but took longer time on medium with or 1 mgL-1 NAA + 5 mgL-1 BAP as well as medium with 1 – 2 mgL-1 NAA only. The same phenomenon was also found on medium with 1 – 2 mgL-1 NAA only. This study also revealed that most of cultured explants regenerated only one shoot, but explants cultured on medium with 1 mgL-1 NAA along with 1 mgL-1 BAP produced 2.33 shoots on the average. The addition of 1 mgL-1 NAA without BAP and 1 mgL-1 BAP without NAA could increase average shoot growth by 1.78 and 1.44, respectively. Keywords: in vitro culture, micropropagation, naphthalene acetic acid, benzyl amino purine.
... The normal concentration range of Zn in plant tissues is reported to be about 25-100 ppm (Campbell 2000;Obreza et al., 2010). Zinc concentration in soils typically ranges between 10-300 ppm (Alloway 2004), and Zn is included in in vitro growth media at concentrations of between 1.5×10 4 -0.9 ppm (Saad and Elshahed, 2012). ...
Reactive oxygen species (ROS) are generated in response to the various environmental stresses, and overproduction of ROS can lead to the lipid peroxidation, which often is monitored by measuring the malondialdehyde (MDA). Nevertheless, the extent of oxidative stress is partly dependent on the total antioxidant capacity (AOC) of plant cells. In many experiments, the assessment of oxidative stress is based on the measurement of ROS only or more commonly individual antioxidants, but a more reliable assessment of oxidative stress requires the simultaneous measurement of ROS, AOC and MDA levels. In this study, the effects of different ZnO concentrations were studied on the oxidant and antioxidant potentials, and also on MDA levels in tobacco plants. Moreover, the significant correlations were considered between the indices, which were evaluated via different methods. Results showed that both Zn deficiency and excess levels led to the increased ROS/H2O2 production, Chl a/b reduction, electrolytes leakage and lipid peroxidation, compared to the control. The inverse conditions were observed for the adequate ZnO amount (1μM) with the highest AOC level. ROS was significantly correlated with H2O2 (P<0.01, R= 0.8) and MDA (P<0.01, R= 0.6-0.8) but not with AOC. Only MDA-PhB method caused a significant correlation (P<0.01, R= -0.7) with the results from AOC methods. Unlike Chla/Chlb, the electrolyte leakage was often significantly correlated with other stress indices. In general, a dependency to the used method was observed for finding the significant correlations among oxidative stress markers, and their simultaneous measurements were also suggested for the better interpretation of data.
... Plant tissue culture media are employed for plant growth and propagation and provide controlled conditions such as necessary inorganic compounds, adequate pH levels and sources of carbon 12,13 . Previous studies investigating the selection of the appropriate basal media for the formation and growth of hairy root cultures (HRCs) and the accumulation of desirable natural products have shown that the ideal medium is dependent on the plant species in question [14][15][16][17][18] . ...
Watercress (Nasturtium officinale) is considered to be one of the most promising plant sources of a variety of glucosinolate compounds. The present study investigated the effect of various types of media on growth and glucosinolate production in the hairy root cultures of watercress. Hairy root cultures are generated using watercress explants infected with Agrobacterium rhizogenes strain R1000. We used six types of media (B5, 1/2 B5, MS, 1/2 MS, SH and 1/2 SH media) in the establishment of hairy root cultures and quantified variations in the production of four glucosinolates (4-hydroxyglucobrassicin, glucobrassicin, 4-methoxyglucobrassicin and gluconasturtiin). Of the six media conditions, MS basal medium increased the accumulation of glucosinolates wherein the contents of glucobrassicin, 4-methoxyglucobrassicin and 4-hydroxyglucobrassicin were 25, 44.88 and 217.33 times higher respectively than those in 1/2 SH medium. Moreover, gluconasturtiin content was 10.55 times higher than that in 1/2 B5 medium. Conversely, SH and 1/2 SH media were the most suitable for hairy root growth. These results suggest an alternative approach for glucosinolate production in watercress hairy root cultures.
... Organogenesis (type and extent) in plant cell cultures is determined by the proportion of auxins to cytokinins [78]. Cytokinins, such as kinetin and BAP, have been proven to promote cell division, shoot proliferation, and shoot morphogenesis and to repress root formation; whereas auxins, such as NAA and dicamba, are usually used to stimulate callus production and cell growth, to initiate shoots and rooting, to induce somatic embryogenesis, and to stimulate growth from shoot apices and shoot stem culture [79]. In this study, complete plantlets (with roots and shoots) were obtained from M1 (containing kinetin) and M2 (containing NAA + BAP) media compositions. ...
In vitro culture offers many advantages for yam germplasm conservation, propagation and international distribution. However, low virus titres in the generated tissues pose a challenge for reliable virus detection, which makes it difficult to ensure that planting material is virus-free. In this study, we evaluated next-generation sequencing (NGS) for virus detection following yam propagation using a robust tissue culture methodology. We detected and assembled the genomes of novel isolates of already characterised viral species of the genera Badnavirus and Potyvirus, confirming the utility of NGS in diagnosing yam viruses and contributing towards the safe distribution of germplasm.
... The nutrition components of medium also affect on callus induction. There are some nutrients only found in WPM medium while they are lacking in MS medium such as calcium nitrate, potassium sulfate and mangan sulfate (Saad and Elshahed, 2012). Those nutrients are likely to be the factor which influence the responses of explants towards the formation of callus (Osman et al., 2016). ...
Sapodilla (Achras zapota) is one type of tropical fruits with high economic value that are easily found in Indonesia. However, the number of productive plants has been decreased and affected the rate of fruit production. The objective of this research is to develop in vitro propagation of Achras zapota through meristem induction to provide great quality seedling. Type of explants (apical and lateral meristem) are cultured on Woody Plant Medium (WPM) and combined with different concentrations of BAP (0 ppm, 2 ppm, 3 ppm, 4 ppm, 5 ppm). Variables observed are shoot, leaf, root (emergence, length, number) and callus formation. The results show that all treatments able to induce shoot and leaf, but only several treatments able to induce root and callus formation. Lateral meristem is an appropriate explant to induce high number of leaves significantly and lateral meristem has the highest rate of multiplication 1.7 with application of BAP 2 ppm. The results can be used as reference to promote shoot multiplication. But further research is needed to determine proper combination to induce shoot multiplication significantly. © 2016 Endang Yuniastuti, Novita Chrisna Wardani and Nandariyah.
... Cell growth and the accumulation of bioactive compounds in plant cell culture are affected by several factors, such as culture media and culture conditions. A plant tissue culture medium generally contains macronutrients, micronutrients, vitamins, amino acids or nitrogen supplements, a carbon source, undefined organic supplements, and growth regulators (Abobkar and Ahmed 2012). The optimum concentration of each nutrient for achieving maximum growth rate and metabolite production varies among different plant species. ...
The optimal culture medium for the production of flavonoid compounds from Orostachys cartilaginea V. N. Boriss. calluses was studied. In callus cultures of O. cartilaginea, the flavonoid monomer content, in decreasing order was: kaempferol-3-O-rutinoside (Kp-3-rut), quercetin 3-O-glucoside (Qc-3-glc), epicatechin gallate (Ecg), kaempferide (Ke), and quercetin (Qc). The results of the uniform design experiment indicated that the production of Qc, Ke, Qc-3-glc, Kp-3-rut, and total flavonoids were satisfactory in callus grown on full salt strength (1) of MS medium supplemented with 3.5 mg L-1 6-benzylaminopurine (BA) and 0.1 mg L-1 1-Naphthaleneacetic acid (NAA). By contrast, only Ecg was found in callus grown on 0.75 MS medium supplemented with 1.5 mg L-1 BA and 0.3 mg L-1 NAA. A phosphate concentration of 1.25 mM in the MS medium favored the production of Qc and Ke, whereas 0.75 mM phosphate was optimal for the production of Ecg, Qc-3-glc, Kp-3-rup, and total flavonoids. The NH4+/NO3- ratios of 30/30 mM in the MS medium promoted Ke, Ecg, Qc-3-glc, Kp-3-rup, and total flavonoid production. However, a NH4+/NO3- ratio of 20/40 mM enhanced Qc production. The effect of sucrose concentrations on the accumulation of different flavonoid monomers was comparatively more regular. The flavonoid content increased as the sucrose concentration increased from 20 g L-1 to 40 g L-1, peaked at 40 g L-1, and decreased at concentrations greater than 40 g L-1. Therefore, 40 g L-1 sucrose was optimal for the production of the five flavonoid monomers and total flavonoids. The present findings demonstrate the possibility of producing flavonoid compounds from O. cartilaginea callus.
... It is frequently used as a cut flower in social and religious ceremonies (Mitra, 1992). It is sold at almost every corner of Dhaka city by the retailers (Hossain, 1995). In Bangladesh, its demand is increasing because of its elegant spike, rich varied colours and long vase life. ...
An investigation was conducted with a view to observing the performance of two gladiolus genotypes during the rainy summer under poly tunnel production system. The genotype GL-027 produced taller plants (73.8 cm) with longer spike (97.2 cm) and longer rachis (51.7 cm) compared to the genotype GL-023. In respect of spike length (97.0 cm), rachis length (54.4 cm), weight of corm per plant (16.0g) and weight of cormel per plant (30.3 g), 1 July planting was better than in 1 May planting. Plants grown under poly tunnel produced flower earlier (82.5 days) compared to plants grown without poly tunnel (84.6 days). Although difference of only two days in flower initiation is of no significance, but plants grown under poly tunnel performed better in respect of number of florets per spike (12.8), number of cormels per plant (8.2), weight of corm per plant (16.0 g) and weight of cormels per plant (30.6 g) than the plants grown without poly tunnel. Keywords: Gladiolus; protected cultivation; rainy season. DOI: http://dx.doi.org/10.3329/bjar.v36i2.9256 BJAR 2011; 36(2): 285-290
... For micropropagation, most widely used culture medium is Murashige and Skoog, 1962 (MS) medium, because most of the plants respond favorably to this medium, contained all the nutrients essential for in vitro plant growth (Kumar and Reddy, 2011). Full strength of salts in media provide good results for numerous species, but in some plants and for specific purposes the reduction of salts level to half or quarter of the full concentration gave better results in in vitro growth (Saad and Elshahed, 2012). Most of the authors studied in vitro cultures of A. montana recommended using half strength MS medium for rhizogenesis (Conchou et al., 1992;Le, 2000;Petrova et al., 2011), however the effect of strength of the MS medium on micropropagation and rooting of the species is not sufficiently Bio Bulletin 2(1): 6-13(2016) (Published by Research Trend, Website: www.biobulletin.com) ...
... Subsequent optimisation improved the celldoubling time from 106 to 50 h and the yield of vanillin from 100 to 1000 mg l À1 (80 g kg À1 on dry weight basis). 62 The addition of Amberlite adsorbents XAD-4 and XAD-7 to the culture medium resulted in excretion of metabolites from cells to medium. XAD-4 effectively adsorbed vanillin, leading to an enhancement in vanillin accumulation. ...
This review deals with the Vanilla plant: history; botanical description; chemistry of vanilla beans; curing of vanilla beans; commercial extraction of vanilla flavour; standard specifications and uses of vanilla flavour. The production of vanillin by both chemical and biotechnological methods is described. The biotechnological production of vanilla flavour metabolites by plant tissue/cell culture, microbial biotransformation and molecular approaches is also presented, together with a discussion on economic and safety considerations.© 2000 Society of Chemical Industry
Various rapid propagation strategies have been discovered, which has facilitated large-scale plant reproduction and cultivar development. These methods, in many plant species, are used to rapidly generate large quantities (900 mini-tubers/m 2) of high-quality propagule (free from contamination) at a relatively low cost in a small space. They are also used for plant preservation. This review article aims to provide potential applications for regeneration and clonal propagation. Plant propagation using advanced agrotechnology, such as aeroponics, is becoming increasingly popular among academics and industrialists. The advancement of asexual aeroponic propagation has been achieved through advancements in monitoring and control systems using IoT and smart sensor technology. New sensor technology systems have gained substantial interest in agriculture in recent years. It is used in agriculture to precisely arrange various operations and objectives while harnessing limited resources with minimal human intervention. Modern intelligent technologies and control systems simplify sensor data collection, making it more efficient than manual data collection, which can be slow and prone to errors. Specific ambient variables like temperature, humidity, light intensity, stock solution concentrations (nutrient water), EC (electrical conductivity), pH values, CO2 content, and atomization parameters (frequency and interval) are collected more effectively through these systems. The use of intelligent technologies provides complete control over the system. When combined with IoT, it aids in boosting crop quality and yield while also lowering production costs and providing data directly to tablets and smartphones in aeroponic propagation systems. It can potentially increase the system's productivity and usefulness compared to the older manual monitoring and operating methods.
Orchids are appreciated worldwide for their unique and beautiful flowers which hold a large commercial and floricultural value. Other than being ornamental, orchids are known to possess medicinal properties as well. Orchidaceae is one of the largest families of flowering plants with ~30,000 species reported worldwide. In India, there are over 1000 species of documented orchids and the Northeast region of India is the home to many such orchids. Many of them are rare and endangered species and belong to Appendix I of CITES (Convention on International Trade in Endangered Species of Wild Fauna and Flora). Their existence is highly threatened due to extensive exploitation and habitat destruction and as such their conservation is of utmost importance. One way to conserve such exotic wild species of orchids is through the intervention of biotechnological methods such as tissue culture. The chapter focuses on the up-to-date progress on the propagation and conservation of rare, endemic and endangered important orchids of the Northeast region of India which are of medicinal values. Phytochemicals that have been identified in many of the orchid species are responsible for their therapeutic properties.KeywordsOrchidaceaeNortheast regionConservationEthnomedicineTissue culturePhytochemicals
This edited book is focusing on the novel and innovative procedures in tissue culture for large-scale production of plantation and horticulture crops. It is bringing out a comprehensive collection of information on commercial-scale tissue culture with the objective of producing high-quality, disease-free and uniform planting material. Developing low-cost commercial tissue culture can be one of the best possible ways to attain the goal of sustainable agriculture. Tissue culture provides a means for rapid clonal propagation of desired cultivars, and a mechanism for somatic hybridization and in vitro selection of novel genotypes. The application of plant tissue culture technology in horticulture and plantation crops provides an efficient method to improve the quality and nutrition of the crops.
This book includes a description of highly efficient, low cost in vitro regeneration protocols of important plantation and horticulture crops with a detailed guideline to establish a commercial plant tissue culture facility including certification, packaging and transportation of plantlets. The book discusses somatic embryogenesis, virus elimination, genetic transformation, protoplast fusion, haploid production, coculture of endophytic fungi, effects of light and ionizing radiation as well as the application of bioreactors.
This book is useful for a wide range of readers such as, academicians, students, research scientists, horticulturists, agriculturists, industrial entrepreneurs, and agro-industry employees.
Somatic embryogenesis has seen many advances. However, many aspects are not fully understood especially on cashew (Anacardium occidentale) despite several studies conducted for the technique improvement. Regenerate in vitro viable embryos through asexual cells (haploid or diploid) is the target of somatic embryogenesis. This process leads to the production of bipolar structures with root/shoot axis well defined. The in vitro culture of cashew is brought out with emphasis on the critical factors that influence the explants response and plantlet regeneration. The recalcitrant nature of cashew has been attributed to abnormal development observed in the calli derived from its explants in some cases and to the limited success recorded up to here in tissue culture of the plant. This review highlights advances, challenges, and future prospects in somatic embryogenesis research of cashew.
The demand for economically important plants, particularly medicinal and horticultural crops has increased drastically. On a large scale, the availability of propagules for the cultivation of these crops remains a challenge since plant parts such as seeds, rhizomes, and corms needed for propagation are often the economically important parts in high demand. For decades, tissue culture techniques have been used to bridge the supply versus demand gap for both medicinal plants and horticultural crops. However, mass propagation of plants in vitro is hampered by the costs associated with such in vitro techniques. In tissue culture protocols, the cost of prepared growing media, amongst other inputs, can be a limitation. Murashige and Skoog (MS), Linsmaier and Skoog (LS), Gamborg (B5), Lloyd and McCown (LM) Woody Plant, Driver and Kuniyuki Woody (DKW), BDS, BABI as well as Nitsch and Nitsch (NN) media have been used in tissue culture as basic media for decades. Main components of these media include micro- and macronutrients, vitamins, amino acids or nitrogen supplements, source (s) of carbon, and undefined organic supplements in some cases, which are solidified by gelling or solidifying agents. To attain cost-effective protocols for mass propagation, media containing different low-cost components can be used as alternatives. However, mass propagation protocols are often species-specific or vary amongst species due to differences in nutrient requirements and plant physiology. Optimization of components is imperative for achieving low-cost high multiplication and survival rates during acclimatization. This chapter aims to explore low-cost media for mass propagation of clonal plants to sustain the agricultural industry and counteract food insecurity.
Explant preference is a key factor for efficient and sustainable plant propagation under in vitro conditions. Plant genotype and structure must be well observed and identified for the best explant which may differ in the axillary bud breakings using terminal buds on stems located above ground or specialized/underground stems such as bulbs scales, base plates of corms, and the shoot tips of suckers. Since plant factory systems are aimed at uniform and cost-effective propagation systems, determination of explant type and culture conditions are the most critical factors for the establishment of shoot multiplication rate. In this chapter, several horticulture plants including house plants (Monstera, Philodendron, Begonia, etc.), and fruit trees (Aronia, banana, walnut, etc.) used in commercial-scale production in plant factories were investigated for the understanding of the nature of explants as per culture conditions. This phenomenon is also highly correlated with effective surface sterilization. Since plant factories rely on an automation system for particular crops, replenishment of starting material in each cloning cycle prevents the emergence of undesirable traits due to the somaclonal variations. This study reports a comparative and in situ analysis of explant choice for the scalable vitro-plant productions.
Strawberry plants grown in vitro are typically stored and maintained on agar containing Murashige and Skoog (MS) media and sucrose as a carbohydrate source. This method of storing strawberry plants in vitro is expensive and time consuming, requiring sub-culturing onto fresh media every 2 to 3 mo. This study aimed to establish the viability of using a substrate alternative as an economical replacement for MS media, for both long-term storage in vitro and ease of transfer ( ex vitro ). A protocol was developed for strawberry in vitro using commercially available sterilized peat pellets (Jiffy-7® pellets), to optimize culture conditions and tissue culture practices. Suitability of the peat substrate was measured by the plant’s overall response to culture in vitro and subsequent health and survival ex vitro following deflasking. Included in this study was a comparison of the use of vented vs non-vented tissue culture vessels for their effect on plant development and survival in vitro and ex vitro . The results show that strawberry plants can be grown and stored under in vitro conditions in vented vessels without sub-culturing for up to 3 yr or more. This equates to an approximately eightfold more efficient technique, significantly decreasing cost of storing and maintaining strawberry plants in vitro . Furthermore, plants grown in the peat substrate did not multiply, and had more established secondary roots than those grown in the conventional MS media.
Buckwheat is a member of a genus of 23 species, where the two most common species are Fagopyrum esculentum (common buckwheat) and Fagopyrum tataricum (Tartary buckwheat). This pseudocereal is a source of micro and macro nutrients, such as gluten-free proteins and amino acids, fatty acids, bioactive compounds, dietary fibre, fagopyrins, vitamins and minerals. It is gaining increasing attention due to its health-promoting properties. Buckwheat is widely susceptible to in vitro conditions which are used to study plantlet regeneration, callus induction, organogenesis, so-matic embryogenesis, and the synthesis of phenolic compounds. This review summarises the development of buckwheat in in vitro culture and describes protocols for the regeneration of plantlets from various explants and differing concentrations of plant growth regulators. It also describes cal-lus induction protocols as well as the role of calli in plantlet regeneration. Protocols for establishing hairy root cultures with the use of Agrobacterium rhizogens are useful in the synthesis of secondary metabolites, as well as protocols used for transgenic plants. The review also focuses on the future prospects of buckwheat in tissue culture and the challenges researchers are addressing.
Quinoa is a pseudocereal which is growing popularity day by day in the scientific world due to its rich nutrient profile and easy cultivation. Several scientists are taking interest in these novel grains in order to study as well as to learn the mechanisms underlying its tolerance to diverse stress conditions. In order to prepare fresh grains for work in the laboratory, it is essential to ensure sterility from contaminating microorganisms so that no interference can be caused during observations. In the field, several millions of microbes come in contact with the grains and other parts of the plant which live either as symbionts, parasites, and even pathogens. These microbes exist in nature within high concentrations in soil and sometimes even help and support the plant growth by establishing symbiotic relationships. Parasites and pathogens, on the other hand, possess the ability to destroy the whole field. It is necessary to take various precautionary measures to prevent their entry in the vegetation through various physical and sometimes chemical and biological practices. This chapter also discusses in depth the composition of two culture media that can be used in laboratory to cultivate this crop which are Murashige and Skoog (MS) media and Gamborg’s B5 media for tissue culture operations, aseptic cultivation, and practices that need to be undertaken for maintenance of plant cell cultures in vitro.
Conventional plant breeding has been going on for hundreds of years and is still commonly used today by artificially mating or cross‐pollinating to increase yields. Plant tissue culture was introduced to facilitate the clonal propagation of horticultural species. Based on the material used, the technology of plant tissue culture may be divided into three domains: whole plant culture, plant organ culture, and protoplast culture. Nutrient media plays an important role in tissue culture and it is very vital for proper and timely growth of cells and their multiplication. Genetic engineering techniques in cereal plants, fruits plants, legumes and vegetable plants have the capacity to become low cost bioreactors to make molecules that in the normal scheme of things would not have been available from plants. Governments are concerned on the safety, access, and profitability of biotech crops, as well as local interests on biodiversity protection and trade competitiveness.
In this study, multiple and rapid propagations of Staurogyne repens (Nees) Kuntze with tissue culture techniques were aimed. The nodal explants of S. repens were cultured for multiple shoot regeneration in culture medium supplemented with different rates of 6-Benzilaminopurin (BAP) and Kinetin (KIN) alone (0.25-1.50 mg/L) or combinations with 0.25 mg/L Indole-3-acetic acid (IAA). Generally, shoot regeneration capabilities of the explants were high due to the growth regulators used. Shoot regeneration frequencies were ranked between 77.77% and 100.00% in BAP-IAA and KIN-IAA applications. In BAP-IAA application, the average number of shoots was recorded between 2.53-16.11 shoots/explant and the highest number of shoots per explant (16.11 shoots/explant) was obtained in culture medium containing 1.25 mg/L BAP. As the BAP level used increased, the number of regenerated shoots increased. In KIN-IAA application, mean shoot counts were recorded as 2.66-10.27 shoots/explant, and maximum shoots per explant (10.27 shoots/explant) was determined in culture medium containing 0.50 mg/L KIN. When shoot lengths were examined, longer shoots were seen in KIN-IAA application compared to BAP-IAA application. The longest shoot was obtained as 2.82 cm in the culture medium fortified with 1.50 mg/L KIN. Regenerated shoots for in vitro rooting were transferred to the growth culture containing different concentrations of IAA and 1-Naphthaleneacetic acid (NAA). The best values in terms of number and length of roots per shoot were determined in a growth culture containing 0.25 mg/L IAA. The rooted shoots were accustomed to external conditions successfully.
One of the methods in sustainable agriculture is transforming crop wastes into beneficial products such as bio-fertilizer, which will improve the soil fertility, preserving the natural sources, and cost-effective practices. Currently, these practices also need to be applied in plant tissue culture technology to enhance an eco-friendly and sustainable resources which are abundant and locally available. To date, the new bio-organic media derived from different types of agricultural wastes, especially from fruit wastes were successfully established. The objective of this study was to determine the efficiency of agricultural waste, i.e. Palm Oil Mill Effluent (POME) through a natural fermentation process as a potential nutrient source to substitute the existing standard and synthetic plant tissue culture media (PTC) such as Murashige and Skoog’s (1962) (MS) Medium, Woody Plant Medium (WPM), N6 (Chu) Medium and Gamborg’s (B-5) Medium, available in the market. 100 mL (v/v) POME solution was added with 100 mL (v/v) molasses and then were fermented naturally for 4 weeks in dark condition at room temperature. The results showed that the nutritional value of the fermented POME solution was higher than the synthetic MS basal salt medium. The new POME PTC medium was successfully formulated as a new bio-plant tissue culture media for in vitro regeneration of Musa acuminata.
Plants have proven to be a beneficial means for uncovering new products having therapeutic interest in the drug augmentation. Human beings uses plant-produced secondary metabolites since from the prehistoric times. Due to high usage of secondary metabolites in diverse marketing sectors, such as pharmaceutical, food, and chemical industries, the demand for the most relevant and accepted method to separate these metabolites from plants is huge. Different extraction techniques have been used to obtain secondary metabolites, and many of these techniques are built on the extracting strength of solvents and the application of mixing and/or heat. In addition to traditional methods, several new methods have been established, but till now none of them are considered as a standard method for elicitation of secondary metabolites. In the late 1960s, plant cell culture technologies were found as a promising tool for both investigating and designing plant secondary metabolites. With the help of cell cultures, phytochemicals are not only produced in adequate quantity, but also discard the existence of intrusive compounds that develops in the field-grown plants. This technology serves advantageous over classical methods. Many approaches have been used to amplify the yield of secondary metabolite manufacture by cultured plant cells. Among these approaches are selecting a plant with immense biosynthetic capacity, acquiring efficacious cell line for growth and production of the concerned metabolite, manipulating culture environment, elicitation, metabolic engineering, and organ culture. Mass cultivation of plant cells is done with the help of different bioreactors. Application of cell culture provides various benefits including the synthesis of secondary metabolites, working in controlled conditions as well as autonomous to soil and climate conditions. Elicitor which may be biotic or abiotic is considered as one of the stress agents to obtain increased amount of secondary metabolites from different parts of the plants. Polysaccharides like chitosans are natural elicitors which are benefitted for plant cell’s immobilization and permeabilization. A new path has been initiated in current years for secondary metabolite production with the help of elicitors in plant tissue culture. The different criteria that influence the production and accumulation of secondary metabolites include elicitor concentrations, exposure time, cell line, nutrient composition, and age or stage of the culture. In a number of plant cell cultures, elicitors have intensified the production of sesquiterpenoid, phytoalexin, terpenoid indole alkaloids, isoflavonoid, phytoalexins, coumarins, etc. Regardless of these efforts of the past few decades, plant cell cultures have led to very little economic successes for the production of esteemed secondary compounds. Thus, the aim of this chapter is to highlight the prospects of plant cell culture to produce secondary metabolites, and also provides an overview on the important approaches used for the secondary metabolite production and their improvement strategies.
In this study, we developed an intuitive remote-controlled manipulator system that was operated using a touch panel display and a mouse. We examined its application possibility to pick out the shoot apical meristem from sprouting potatoes. The shoot apical meristem is not easily affected by plant viruses. Therefore, we can obtain a virus-free seedling that has a high production efficiency by culturing this part. This technique is called “mericlone” and is known widely in agriculture; however, it is tedious because of the precision work involved with a microscope. Thus, our system not only reduces the burden on the technician but also has high precision control and saves space. In this study, using the loop-mediated isothermal amplification method, we confirmed that we could get the potato meristems that do not contain Potato Virus X. The newly designed primer set can achieve results more rapidly, more easily, and with higher sensitivity than other virus detection methods, thus indicating the possibility of industrial implementation.
The variety MD2 of pineapple (Ananas comosus L.) was used in this study. This variety is highly demanded in the international market and known for possessing harvest quality with aroma, high sugar content (14% Brix), vitamins and longer shelf life. However, shortage of planting material has limited the production in Malaysia. In vitro mass propagation using the direct and indirect shoot proliferation techniques was tested on variety MD2. The plantlets were successfully initiated from sucker on solid MS basal medium containing 30 g/L sucrose, 0.1 g/L Myo-inositol and 3 mg/L BAP after one month of culture. The highest direct shoot tips regeneration was obtained on solid MS medium when added with 30 g/L sucrose, 0.1 g/L Myo-inositol, 3 mg/L BAP and 1 mg/L NAA. Indirect shoot regeneration was obtained on medium containing 3 mg/L Zeatin after one month of culture. In average, 10 shoots were regenerated from approximately 1 gram of calli. The techniques can produce 100-200 number of plantlet within 4 to 6 months of culture, and ready for planting after 7 months of culture.
Fungi, bacteria and yeast contamination causes several lost in the commercial micro dissemination of vegetal species and in research works. The control of in vitro contamination is essential to increase the micro dissemination efficiency. This work evaluated bacteristatic and/or bactericidal action of PPM (Plant Preservative Mixture) to control the contamination with Pseudomonas, and the effect on the "in vitro" sugarcane growth. The bacterium was isolated from contaminated vegetal material. PPM concentrations were: T1 (0.06); T2 (0.125); T3 (0.250); T4 (0.500); T5 (0.750 ); T6 (1) and T7 (0 mL/L) incorporated to the MS (1962) culture medium. The bacterium was sown in tubes with 5 replications. The bacterial growth was evaluated doing microscopic extended. Using inhibitory concentrations of the bacterial growth and a control without PPM, it was evaluated the effect on the stools growth. Six stools of 2-4 cm long were put in each bottle with 10 replications. Number; length and fresh weight of stools were evaluated. T3, T4, T5 and T6 inhibited the bacterial growth with bactericidal action. There was no inhibitory effects of PPM on the length, fresh weight and number of stools. T5 presented major uniformity in the development of the stools.
This article deals with the theories, methodology and practices of in vitro virus elimination. Viruses may be eradicated from infected stocks through meristem or callus cultures due to the low titre, or even absence, of viruses in plant meristems and what might be called the viral elimination power of in vitro incubation. The season and location of the buds obtained, as well as the explants' size, are important factors in virus elimination through meristem cultures. Heat treatment and high phytohormone concentrations frequently improve the results. Virus-free tissue sectors may arise from calli of infected plants. From such sectors viruse-free plants may regenerate. Nevertheless, the frequently occurring chromosome abnormalities in callus cultures may become a strong drawback for regenerating virus-free plants from callus. Rigid virus-testing procedures must be applied to reveal the virus-free status of the regenerated plants. The commonly used testing methods are briefly described. Methods employed for rapid in vitro mass-cloning of virus-free plants include: multiple shoot formation, in vitro layering, adventitious organogenesis, and in vitro storage organ induction. The resulting virus-free clones frequently exhibit an improved yield and quality. However, some of the mild virus strains display a cross-protection effect. The removal of these viruses through tissue cultures may result in an outbreak of more severe diseases.
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