Large-scale Multiplication Research Articles

Effect of Organic Substrates on Macro-propagation of Banana under Island Conditions

Banana is an important tropical fruit crop grown globally and is the world’s most important agricultural food commodity. Plantation crops take a major share of cultivated area in Andaman and Nicobar Islands. Banana is one of the important fruit crops that is highly suitable to grow as intercrop in plantation based cropping system. Local varieties of banana like Korangi, Mitta Champa, Khatta Champa and Cheenakela are cultivated by the farmers traditionally to meet the local demand. However, in recent past growing commercial varieties of banana has become popular in the Island due to consumer demand. Traditionally banana is propagated by suckers and wide variability was observed in sucker production among the commercial varieties. Due to this large scale multiplication of banana through suckers could not meet the planting material demand in the Island. Hence, a study was undertaken during 2020-2022 to standardize the macro-propagation technology using different organic substrates in the commercial variety ‘Poovan’. Nine different organic substrate combinations were used in the present study to enhance the efficacy of lateral bud development and plantlet production. Suckers weighing 1.0 to 1.5 kg were used as the propagule. Among all the organic media combinations tested, significant differences were observed in terms of plantlet production, root and shoot growth. The treatment T8 (Coircompost: sawdust: FYM: vermicompost (1:1:1:1) with Arka microbial consortia @10 kg) produced maximum number of primary (5.60) and secondary buds (12.07) during primary and secondary capitation of decorticated mother rhizome. Early bud emergence (11.5 days), maximum number of plantlets (12-13 per mother rhizome), maximum shoot length (46.43 cm), maximum shoot girth (3.03 cm) and maximum number of leaves (4.50) were also observed in the treatment T 8. Minimum number of days was taken (52.70 days) for plantlet separation during secondary capitation in the same treatment T 8 suggesting that the treatment combination (Coircompost: sawdust: FYM: vermicompost (1:1:1:1) with Arka microbial consortia @10 kg) was effective for macro-propagation of banana Cv. Poovan under Island ecosystem.

Fertigation with Nitrogen and Phosphorus Improve Plant Growth, Root Growth, and Buddability of Containerized Citrus Nursery Plants

ABSTRACT Large-scale multiplication of quality planting material is needed to meet the huge demand of planting material in fruit crops. Little information is available on the effect of fertigation on rootstock growth, budding success, and buddling growth in containerized citrus nursery. The present studies were undertaken to study the effect of eight fertigation treatments viz. control (N0P0), N60, N80, N100, N0P100, N60P100, N80P100, and N100P100 on the growth of rough lemon rootstock and subsequently budded three citrus varieties. Among the treatments, N60P100 significantly improved seedling height, diameter, leaf number, root growth, leaf area in rough lemon, Blood Red orange, Kinnow mandarin, and Daisy mandarin over the control. The stomatal conductance was higher with treatments N60, N80, N60P100, and N80P100. While higher photosynthetic rates were observed with N60P100 and N80P100, N60P100 also improved the budding success in Blood Red, Kinnow, and Daisy. Fertigation treatments increased the nutrient concentration in leaves of all the varieties. The principal component analysis revealed that plant height and diameter were positively correlated with root length, root diameter, root number, photosynthetic rate, stomatal conductance, and chlorophyll content in rough lemon. In all the varieties, the sprout length and diameter showed a positive correlation with leaf area, leaf number, root length, root diameter, fibrous root number, photosynthesis rate, stomatal conductance, and chlorophyll content. Among different treatments, N60P100 was the best treatment in improving the shoot growth, root growth, and physiological parameters of rough lemon rootstock. Besides, it also improved budding success and buddling growth in all three citrus varieties.

Micropropagation of Zuccagnia punctata, an argentine medicinal plant, by means of “in vitro” technology to promove its conservation and the sustainable production of bioactive compounds

Zuccagnia punctata Cav. (“jarilla poposa”) is a monotypic argentine native medicinal plant used by indigenous communities for the treatment of several pathologies. The pharmacological properties have been scientifically validated and its potential use in phytocosmetic and pharmaceutic industry as antiaged, hypoglycemic, anti-inflammatory, antitumoral, antioxidant, antibacterial and antimicotic has been previously demonstrated. However, the species has a low propagation rate in natural conditions and has been included in the preliminary red list of endangered plants in category 3. Therefore, it is necessary to develop methodologies leading to its conservation and propagation to achieve its sustainable use. The objective of this work is to perform a micropropagation protocol for Z. punctata species for the sustainable use and conservation of this species. Furthermore, the study evaluated also the impact of “in vitro” propagation on the content of bioactive compounds of ethanolic extracts obtained from Z. punctata plants obtained “in vitro” with collected wild plants. Seeds have been used as mother material. The seed germination to obtain explants were performed on Murashige and Skoog medium (MS) without plant growth regulators (PGRs) containing sucrose, agar, and ascorbic acid. The same medium was employed for the multiplication of shoots. “In vitro” rooting was achieved in liquid and semisolid medium, and a higher yield was obtained in the latter. The plants were transferred into commercial substrate and acclimatized in a greenhouse. The micropropagated plants (MP) had a high content of total phenolic compounds similar to those obtained from wild adult plants (WAP) collected in the Monte Desert. The flavonoid content of MP was higher than that of WAP. The main chalcones identified in WAP, 2′,4′ dihydroxychalcone (DHC) and 2′,4′-dihydroxy-3′-methoxychalcone (DHMC) were also present in the MP with a rate 1/1, and 1/3 in WAP. The extracts obtained from the MP showed antioxidant, antiinflammatory and antimicrobial activity, similar to WAP. The “in vitro” propagation and acclimatization of Z. punctata constitutes a fundamental tool to achieve large-scale multiplication of this species in the future and the sustainable production of bioactive compounds with commercial applications. Reimplantation in the natural habitat could prevent the loss of this medicinal genetic resource.

Mass Production of Arbuscular Mycorrhizal Fungi on the Sorghum Plants Inoculated with Burkholderia arboris Using Soybean Mill Waste and Vermicompost-Amended Soil-Sand Substrate.

Arbuscular mycorrhizal (AM) fungi are being used as a new generation of biofertilizers to increase plant growth by improving plant nutrition and bio-protection. However, because of the obligatory nature of the plant host, large-scale multiplication of AM propagules is challenging, which limits its applicability. This study evaluates the ability of Burkholderia arboris to increase AM production in soybean mill waste and vermicompost amended by soil-sand mixture planted with sorghum as a host plant. The experiment was conducted in a nursery using a completely randomized design with four inoculation treatments (B. arboris, AM fungi, B. arboris + AM fungi, and control) under sterilized and unsterilized conditions. AM production was investigated microscopically (spore density and root colonization), and biochemically (AM-specific lipid biomarker, 16:1ω5cis derived from neutral lipid fatty acid (NLFA), and phospholipid fatty acid (PLFA) fractions from both soil and roots). Integrating B. arboris with AM fungi in organically amended pots was found to increase AM fungal production by 62.16 spores g-1 soil and root colonization by 80.85%. Biochemical parameters also increased with B. arboris inoculation: 5.49nmol PLFA g-1 soil and 692.68nmol PLFA g-1 root and 36.72nmol NLFA g-1 soil and 3147.57nmol NLFA g-1 root. Co-inoculation also increased glomalin-related soil protein and root biomass. Principal component analysis (PCA) further supported the higher contribution of B. arboris to AM fungi production under unsterilized conditions. In conclusion, inoculation of AM plant host seeds with B. arboris prior to sowing into organic potting mix could be a promising and cost-effective approach for increasing AM inoculum density for commercial production. Furthermore, efforts need to be made for up-scaling the AM production with different plant hosts and soil-substrate types.

A Review on Plant Tissue Culture

Plant tissue and cell culture involve the aseptic cultivation and propagation of plant cells, tissues, and organs in a meticulously controlled laboratory setting. This innovative method harnesses the potential of nutrient-rich media to efficiently replicate plant cells on a large scale, resulting in the rapid production of mature and disease-free plants. The cornerstone of commercial technology in this field is micropropagation, a process that achieves swift proliferation from minute plant cuttings, axillary buds, and, to a limited extent, from somatic embryos and cell clumps in suspension cultures and bioreactors. Micropropagation, a pivotal aspect of plant tissue and cell culture, holds immense value in generating high-quality and consistent planting materials. These materials find applications across diverse fields, including molecular genetic engineering, plant breeding, horticulture production, and environmental preservation. The process of micropopagation unfolds through several distinct stages, including propagation, subculture of explants for proliferation, shooting and rooting, and hardening. These stages collectively form a universal framework for large-scale multiplication of plants. This technique plays a critical role in overcoming the limitations of traditional plant propagation methods. By allowing for controlled and accelerated growth in a laboratory environment, plant tissue and cell culture contribute to the efficient production of disease-resistant and genetically uniform plant materials. This not only supports advancements in various scientific disciplines but also addresses practical needs in agriculture, horticulture, and environmental conservation. The continuous refinement and application of plant tissue and cell culture methods underscore their significance in meeting the growing demands for sustainable and high-quality plant materials across diverse sectors.

Karatsubha Multiplier by using Verilog

The Karatsuba multiplier is a widely-used algorithm that aims to efficiently multiply large numbers. Traditional multiplication algorithms have a time complexity of O(n^2), where n represents the number of digits in the input numbers. In contrast, the Karatsuba multiplier achieves a faster multiplication process by recursively splitting the numbers into smaller parts and performing intermediate multiplications using only three multiplications instead of four. This Modified architecture saves the 14.9% computation time and it consumes 45.5% less slices than existing Karatsuba multiplier. The proposed architecture has been simulated and synthesized by Xilinx vivado design suite for Spartan & Vertex device family. The new architecture is simple & easy. It emphasizes the significance of the Karatsuba multiplier in improving computational efficiency and highlights its implementation in different domains to accelerate large-scale multiplication tasks. Proposed hardware was implemented on different FPGA devices for various operand sizes, and performance parameters were determined. Comparing to state-of-theart works, the proposed method resulted in a lower combinational delay and area-delay product indicating the efficiency of design. In this we use two 32 bit inputs and produced the 64 bit as the output. In Spartan3E FPGA device family, Modified Karatsuba Algorithm (MKA) is 26.5% faster than Karatsuba Algorithm (KA).It consumes 61.7% less slices than existing KA based Convolution

Double phase culture system mediated enhanced protocol for shoot proliferation of Vanilla andamanica Rolfe. - an endemic wild relative of commercial Vanilla from the Andaman and Nicobar Islands

A liquid overlay culture system in micropropagation improved shoot proliferation in many species. In the present investigation, we have developed an enhanced shoot proliferation protocol using the double phase culture system (DPS) for Vanilla andamanica, an endemic species of the Andaman and Nicobar Islands recognized under the vulnerable category by the IUCN. In vitro generated nodal explants were used for shoot proliferation experiments and were tested in Murashige and Skoog medium augmented with various cytokinins (BAP, KIN, 2iP) and auxins (IBA, NAA, IAA) to produce a maximum of 3.24 ± 0.08 shoots per explant in 5 µM BAP and 5.23±0.67 roots per shoot in basal MS medium. A remarkable increase in shoot regeneration was observed when the nodal explant was cultured on DPS system with optimum cytokinin (5 µM BAP). On evaluation of the influence of DPS and conventional single-phase systems (SPS), (solid medium) exhibits an improved multiplication rate on a DPS with optimal BAP (5µM) with an average of 8.66 ± 0.17 shoots per explant, which represents a 2-fold increase over the rate of SPS + 5 µM BAP. The plantlets were rooted at 100% frequency in half-strength MS medium devoid of auxins and acclimatized with 100% success. A double-phase mediated enhanced shoot proliferating procedure could be employed for large scale multiplication and commercial breeding trials for better results with regard to V. andamanica.

Induction of organogenesis and callogenesis in Limbarda crithmoïdes L. (Asteraceae) explants cultured on MS media supplemented with various concentrations of Na+ and K+

The present work describes an efficient micropropagation and callus induction protocol of Limbarda crithmoïdes L. (Asteraceae), a halophyte species with medicinal and horticultural interests. The objective was to identify the culture media that support the best organogenesis and callogenic expressions of four types of aerial explants; nodal and internodal segments and leaf portions, by varying the mineral composition of twenty culture media by adding increasing concentrations of Na+ (from 0-100 mM) and K+ (from 0-50 mM). After two months of culture, parameters relating to the various expressions of organogenesis and callogenesis were measured. Results showed that the K+ and Na+ interactions affect the explants development. The combination of high concentrations of Na+ (50, 100 mM) and K+ (50 mM) ions allowed the most important regeneration of the axillary shoots (70.6-100%), root neoformation (82.6-96.1%) and callus induction (76.3-100%). Rooted plantlets with well-developed axillary shoots have been successfully acclimatized with a 93% of survival rate. The selected media would allow a large-scale multiplication of this medicinal species, without adding exogenous phytohormones, and could be used for the micropropagation of other threatened halophyte species or for the production of callus; rich in secondary metabolites.

In vitro regeneration through organogenesis in Costus igneus–an important herbal insulin plant

Costus igneus is an important high-valued herbal insulin plant. An efficient protocol for in vitro plant regeneration through organogenesis from leaf and nodal explants was standardized. Explants were incubated in Murashige and Skoog (MS) medium amended with diverse levels of plant growth regulators (PGRs). Optimum frequency (68.25%) of adventitious shoot regeneration was obtained from nodal segments cultured onto MS medium containing 2.0 mg/l of 6-benzylaminopurine (BAP) with an average number of 5.0 shoots per explant. Further, MS medium containing 1.5 mg/l 2, 4- dichlorophenoxyacetic acid (2, 4-D) and 1.0 mg/l Thidiazuron (TDZ) produced a higher frequency of callus induction (55.22%) from leaf explants. Optimum shoot regeneration response (66.0%) through indirect organogenesis was observed from leaf-derived callus on MS medium fortified with BAP (2.0 mg/l) and kinetin (0.5 mg/l) with 6 shoots per callus with mean shoot length of 8.0 cm. The shootlets obtained from node and leaf cultures were rooted with a frequency of (63.52%) on MS medium with 1.0 mg/l indole-3-acetic acid (IAA). Plantlets obtained through organogenesis were successfully acclimatized in the greenhouse and field conditions. There were no significant differences among frequency of somatic plants regenerated through either direct or indirect organogenesis. The present study reports on successful plant regeneration through direct and indirect organogenesis in C. igneus. The regeneration protocols discussed here can be used effectively for large-scale multiplication, germplasm survival, pharmacological and genetic manipulation research.

Fruit Crop Improvement with Genome Editing, In Vitro and Transgenic Approaches

Fruit species contribute to nutritional and health security by providing micronutrients, antioxidants, and bioactive phytoconstituents, and hence fruit-based products are becoming functional foods presently and for the future. Although conventional breeding methods have yielded improved varieties having fruit quality, aroma, antioxidants, yield, and nutritional traits, the threat of climate change and need for improvement in several other traits such as biotic and abiotic stress tolerance and higher nutritional quality has demanded complementary novel strategies. Biotechnological research in fruit crops has offered immense scope for large-scale multiplication of elite clones, in vitro, mutagenesis, and genetic transformation. Advanced molecular methods, such as genome-wide association studies (GWAS), QTLomics, genomic selection for the development of novel germplasm having functional traits for agronomic and nutritional quality, and enrichment of bioactive constituents through metabolic pathway engineering and development of novel products, are now paving the way for trait-based improvement for developing genetically superior varieties in fruit plant species for enhanced nutritional quality and agronomic performance. In this article, we highlight the applications of in vitro and molecular breeding approaches for use in fruit breeding.

Influence of sodium salicylate on adventitious organogenesis of a commercial cucumber cultivar

Cucumber is an important vegetable crop belonging to the Cucurbitaceae family. An optimized regeneration system is required to address different biotechnological approaches in this species, such as large-scale multiplication, as well as genome editing and genetic transformation techniques. The aim of this study was to investigate the effect of different concentrations of the phenolic compound sodium salicylate (NaSA), a salicylic acid derivative, on in vitro callus and shoot regeneration of a commercial cultivar of cucumber. Four-day-old cotyledon explants from the slicing cultivar 'Marketer' were employed. Murashige and Skoog (MS)-derived shoot induction medium containing 0.5 mg L-1 indole-3-acetic acid (IAA) and 2.5 mg L-1 6-benzylaminopurine (BAP) was supplemented with NaSA. Frequency and shoot number were enhanced by 1.5-fold with NaSA at 0.1 µM. Higher salicylate levels led to increased callus formation and decreased shoot regeneration. The application of sodium salicylate at a specific concentration showed a positive trend in in vitro adventitious organogenesis of a commercial cucumber cultivar. Some probable mechanisms that may underlie the beneficial effects of salicylic acid/salicylates on in vitro regeneration were also discussed.

In vitro asymbiotic seed germination, protocorm formation, and plantlet development of Orchis simia Lam.: A threatened terrestrial orchid species

• Seed germination and plantlet growth were impacted by organic constituents and nitrogen sources. • Individual application of organic nitrogen compounds led to higher germination efficiencies. • The present protocol could be used for large-scale in vitro micropropagation of terrestrial orchids. The present study examined the seed morphometric and viability, asymbiotic seed germination, protocorm formation, and plantlet growth in Orchis simia Lam., an at-risk Euro-Mediterranean terrestrial orchid species. Furthermore, tissue histology pattern in vitro grown plantlets and field-established was compared. After one month, shoot and root primordia started to grow on developing protocorms and small plantlets became visible after 2–3 subculturing. Asymbiotic in vitro seed germination and plantlet growth attributes were significantly ( P <0.05) impacted by both type of main organic constituent and nitrogen sources. The seeds sown on media supplemented with pineapple juice (PJ) and casein hydrolysate (CH) had the highest germination percentage. In terms of required days to see the first signs of germination, CW+AV treatment brings about the fastest germination (6.8 ± 0.20 days). Protocorms grown on the media comprising PJ in combination with Aminoven (AV) or casein hydrolysate (CH) or Pep showed higher dimensions and therefore higher weight (% 450 increase) compared to protocorms grown on other media. Altogether, treatments with higher germination percentage, faster germination, and bigger protocorms finally resulted in better plantlet growth. Acclimatization of in vitro grown plantlets occurred by converting scattered vascular tubes into fully developed xylem and phloem vessels facilitating sap flow. Taking into account the fragmentation of O. simia populations in the Middle East and the Euro-Mediterranean basin mainly due to tuber overharvesting and climate change, the present procedure could be used for large-scale multiplication of in vitro raised seedlings to re-introduce into endangered regions.

A comprehensive review of Quercus semecarpifolia Sm.: An ecologically and commercially important Himalayan tree

Himalayan mountain forests have been a potential candidate for the investigation of perturbations due to the complex geography in which they sustain and the sensitivity of the species toward human disturbance and climate change. Among various tree species, brown oak (Quercus semecarpifolia), a very important component of the Himalayan mountains, has been identified as a keystone species due to its substantial economic and ecological benefits. Maintenance of microclimate and suitable habitats with a rich source of natural resources makes Q. semecarpifolia the most preferred forest for luxuriant growth of ground flora, shelter for fauna, and multipurpose uses by the local people. In a climax community, it plays a critical role in environmental balance both at the local and regional levels. Unfortunately, it has become one of the most overexploited tree species of the Himalayan region over the last few decades due to its high demand for dry season fodder and firewood. The wide range of seedling distribution 348–4,663 individuals ha–1 is evidence of the disturbance accompanied by poor regeneration in Q. semecarpifolia forests. Moreover, litter accumulation and grass cover adversely affect seed germination. The ecological cost of oak forest degradation is perhaps more important and damage is irreversible. Thus, continuous demand and extensive threats accompanied by poor regeneration have drawn the attention of stakeholders to conserve this species. However, propagation protocol, especially the pre-sowing treatment of the species, has not been impressive for large-scale multiplication. This review is comprehensive information on distribution, phenology, regeneration pattern, human threat, conservation approaches, and management of Q. semecarpifolia in the Himalayan region.

Micropropagation, encapsulation, physiological, and genetic homogeneity assessment in Casuarina equisetifolia.

Casuarina equisetifolia is an important tree of the forest, cultivated in tropical and subtropical regions, providing fuelwood, land reclamation, dune stabilization, paper production, and nitrogen fixation. We have developed a systematic in vitro propagation protocol in C. equisetifolia using nodal segments (NS). Murashige and Skoog (MS) medium augmented with BA (5.0 μM) and NAA (0.5 μM) gave rise to a maximum of 32.00 ± 0.31 shoots per explant (S/E) with shoot length (SL) of 3.94 ± 0.02 cm, and a maximum of 70% regeneration potential (RP) was recorded after 8 weeks of post inoculation. For root induction, in vitro derived shoots were transferred to the nutrient medium consisting of a half-strength (½) MS medium augmented with 2.5 μM NAA, which produced a maximum of 12.68 ± 0.33 roots/shoot (R/S) with 3.04 ± 0.50 cm root length (RL) in 60% of culture after 6 weeks. Micropropagated plants with healthy shoots and roots were successfully acclimatized in vermicompost + garden soil + sand (1:2:1) and a maximum survival percentage of 95.1% was recorded. NS was taken from a 6-weeks-old in vitro derived plant of C. equisetifolia for synthetic seed production, and it was reported that CaCl2 · 2H2O (100 mM) + Na2-alginate (4%) resulted in clear and uniform beads. Furthermore, the maximum conversion of synthetic seeds into plantlets occurred over a period of 4 weeks of storage at 4°C. Scanning Electron Microscopy (SEM) revealed the formation of direct shoot buds without any intermediate callus formation. In addition, the chlorophyll and carotenoid contents of the direct regenerated and mother plant were compared. Similarly, RAPD and ISSR primers were used for genetic homogeneity assessment of the direct regenerated plants, where a total of 18 and 19, respectively, clear and reproducible bands with 100% monomorphism were recorded. The developed micropropagation protocol can certainly be used for large-scale multiplication and germplasm preservation of C. equisetifolia. It will also help in meeting the growing demands of C. equisetifolia in the forest industry.

Standardization of seed, rootstock and plant standards of lasoda (Cordia myxa L.) for commercial orcharding in India

Aim: The present study was conducted to develop the planting material standards of lasoda cultivar ‘Thar Bold’ for commercial orcharding in India. Methodology: The ripe fruits at pink colour appearance from mother tree of lasoda ‘Thar Bold’ were collected from Experimental Block-I after 75+5 days after fruit set during last week of May- first week of June months for raising of seedlings. Fruits and seeds were categorized based on size and weight and accordingly seeds sown in field bed and root trainers. At two true leaf stage, the seedlings were shifted in polybags for working out seed and rootstock standards. Further, to develop the planting material standards, the scion buds of current season’s growth were collected and budded during third week of May, June and July. The standards were prepared on the basis of all possible traits of the interest. Results: Under the present study, planting material standards of lasoda were developed with following prerequisites: ripened fresh fruits should have diameter &gt; 1.5 cm and fruit weight &gt; 6 g with seed germination &gt; 60 %; buddable seedlings/ rootstock of 9-12 months should have about 35-40 cm height and 0.8-1.0 cm girth; budding should be done during May-June months; at 90 days of budding, plants with sprouts length ≥ 50 cm and diameter ≥ 0.5 cm should be good for field planting in the same season. Interpretation: The developed standards of lasoda would be highly useful for conservation of elite type and large scale multiplication of quality planting material for commercial orcharding in the country.

Acceleration of micropropagation procedure of Bambusa nutans: A commercially important bamboo species

The present communication described an accelerated, efficient and reproducible in vitro propagation procedure for large-scale multiplication of Bambusa nutans Roxb. The nodal segments (12–18 mm) collected from secondary branches of superior clump in the month of April reported enhanced multiple shoot formation (7-8) when implanted on Murashige &amp; Skoog semi solid medium supplemented with 10 µM BA (6-Benzylaminopurine). These axillary shoot cultures pass through ten subculture cycle on liquid medium of 10 µM BA. The cluster of two axillary shoots transferred to liquid MS medium containing 10 µM BA+ 7.0 µM NAA (a-Naphthaleneacetic acid) produced a maximum of 14.5 multiple shoots after 20 days of inoculation. Rooting of 100 % of shoots was achieved in excised propagules when transferred to ½ MS medium supplemented with 30 µM NAA. A two-step hardening procedure of in vitro raised plants ensured 100 % survival. Macroproliferation was carried out in hardened plants after 4 months registered threefold number of plants in polythene bags. The field transferred plants were reported with 100 % survival with prolific new shoot formation and growth.

In vitro Multiplication of Kaempferia Galanga L.- An Important Medicinal Plant

An efficient in vitro propagation protocol was standardized in K. galanga, wherein shoot cultures were raised from rhizome with axillary bud explants in Murashige and Skoog (MS) medium supplemented with different hormonal regimes. The best medium identified for shoot initiation as well as multiplication was MS supplemented with 2 mg/l BAP and 1 mg/l Kin. The high rates of rooting in Half MS media supplemented with 1 mg l−1 IBA and successful transfer to polybags make this production system useful for ex-situ conservation and large scale multiplication of kacholam.

Indirect in vitro Regeneration of the Medicinal Plant, Aspilia africana, and Histological Assessment at Different Developmental Stages.

The medicinal plant, Aspilia africana, has been traditionally used in several African countries to treat many diseases such as tuberculosis, cough, inflammation, malaria, osteoporosis, and diabetes. In this study, we developed a protocol for in vitro propagation of A. africana using indirect shoot organogenesis from leaf and root explants of in vitro-grown seedlings and assessed the tissues at different developmental stages. The highest callus induction (91.9 ± 2.96%) from leaf explants was in the Murashige and Skoog (MS) medium augmented with 1.0 mg/L 6-Benzylaminopurine (BAP) and 1.0 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D) while from root explants, the highest callus induction (92.6 ± 2.80%) was in the same plant tissue culture medium augmented with 0.5 mg/L BAP and 1.0 mg/L 2,4-D. The best shoot regeneration capacity from leaf-derived calli (i.e., 80.0 ± 6.23% regeneration percentage and 12.0 ± 6.23 shoots per callus) was obtained in medium augmented with 1.0 mg/L BAP and 0.05 mg/L α-Naphthaleneacetic acid (NAA); the best regeneration capacity for root-derived calli (i.e., 86.7 ± 6.24% shoot regeneration percentage and 14.7 ± 1.11 shoots per callus) was obtained in the MS medium augmented with 1.0 mg/L BAP, 0.05 mg/L NAA, and 0.1 mg/L Thidiazuron (TDZ). Regenerated plantlets developed a robust root system in 1/2 MS medium augmented with 0.1 mg/L NAA and had a survival rate of 93.6% at acclimatization. The in vitro regenerated stem tissue was fully differentiated, while the young leaf tissue consisted of largely unorganized and poorly differentiated cells with large intercellular airspaces typical of in vitro leaf tissues. Our study established a protocol for the indirect regeneration of A. africana and offers a basis for its domestication, large-scale multiplication, and germplasm preservation. To the best of our knowledge, this is the first study to develop an indirect regeneration protocol for A. africana and conduct anatomical assessment through the different stages of development from callus to a fully developed plantlet.

Genetic diversity among genotypes of Uncaria guianensis (Aubl.) J.F. Gmel. maintained in an in vitro germplasm bank.

The online version contains supplementary material available at 10.1007/s13205-021-03016-y.

Optimization of Callogenesis/Caulogenesis Induction Protocol in Saffron Plant (Crocus sativus L.) Using Response Surface Methodology

The Crocus sativus, an endangered medicinal and aromatic plant in Morocco, has a low propagation rate in natural conditions and, therefore, an efficient method for in vitro propagation is required. This study investigated the effects of various hormones on the induction of callogenesis and callogenesis in C. sativus corms using the Box-Behnken experimental design. The best shoot formation was obtained with Murashige and Skoog fortified with 3 mg/L 6-Benzylaminopurine. On the other hand, callus formation was obtained with 3 mg/L 1-Naphthaleneacetic Acid or 3 mg/L 2,4-Dichlorophenoxyacetic Acid. However, a combination of 3 mg/L 6-Benzylaminopurine, 1.056 mg/L Indole Butyric Acid, and 3 mg/L 2,4-Dichlorophenoxyacetic Acid allows 50% caulogenesis and 60% callogenesis. The in vitro regeneration system could be utilized for both conservation and largescale multiplication of Crocus sativus corms.