Propagation Protocol Research Articles

In vitro development of gametophytes and sporophytes of critically endangered fern Christella kendujharensis S.K. Behera & S.K. Barik (Thelypteridaceae)

AbstractChristella kendujharensis S.K. Behera & S.K. Barik (family: Thelypteridaceae) is a critically endangered fern endemic to India. We developed its mass propagation protocol through in vitro spore culture in a liquid suspended medium for germination of gametophytes that produced sporophytes subsequently. The spores were monolete, reniform (mean length 36.49 ± 3.6 × breadth 25.67 ± 4.2 μm) to spheroidal with fimbriate and broad‐winged perispore. The maximum germination was in the Parker's and Thompson's (P&TV) medium with 0.1 mg L−1 thiamin, 0.1 mg L−1 riboflavin, 0.1 mg L−1 biotin, 0.1 mg L−1 pyridoxin, 0.5 mg L−1 nicotinic acid, 100 mg L−1 myoinositol, 0.01 mg L−1 folic acid, and 3 mg L−1 glycine. Spore germination was Vittaria type and gametophyte development was Aspidium type. The cordate gametophyte developed archegonia on the 38th day and antheridia on the 42nd day of spore sowing. Both isolate and composite cultures developed sporophytes indicating the presence of intra‐gametophytic selfing. Based on the maximum number of regenerated gametophytes, two combinations, that is, (i) 2 mg L−1 kinetin, and 0.01 mg L−1 2,4‐dichlorophenoxyacetic acid, and (ii) 0.1 mg L−1 naphthaleneacetic acid and 2 mg L−1 kinetin in P&TV media were recommended for mass propagation of C. kendujharensis.

Tissue culture technology for sumac (Rhus coriaria) propagation

The cultivation of Rhus coriaria, after a long period of abandonment, is spreading again, thanks to the increasing interest in natural products of which this plant is rich. Traditional sumac propagation, difficult and slow, does not guarantee to cover the plant request for new plantings; in vitro tissue culture can be a valid alternative to supply, in a short time and independently of the season, many plants. In this study, three in vitro tissue culture procedures and several auxin/cytokinin combinations were tested in order to identify a successful protocol for the mass propagation of sumac. A satisfying level of sprouts development was obtained from nodal explant culture and from encapsulated microcuttings, while a scarce response was given by in vitro-cultured petioles. None of the methods tested resulted in adventitious root formation; it will, therefore, be necessary to test further auxin/cytokinin combinations. To the best of our knowledge, this is the first study investigating in vitro sumac propagation, using the encapsulation and petiole culture methods. Results are encouraging and set the basis for further research to improve proliferation efficiency and to induce shoots to root.

Mass Propagation of Dendrocalamus Giganteus Wall. Ex Munro Through in Vitro Culture

The present experiment was conducted to establish an efficient protocol for mass propagation of Dendrocalamus giganteus Wall. ex Munro through in vitro culture using nodal segments as explants. The highest percentage of shoot induction (86.67 %) was obtained in the liquid MS medium supplemented with 1.5 mg/l BAP and 1.0 mg/l TDZ with maximum number of shoots (3.91 ± 0.87) per explant. The highest number and length of shoots were 22.83 ± 1.30 shoots/culture and 7.71 ± 0.96 cm, respectively induced on agitated liquid MS medium supplemented with 2.0 mg/l BAP and 1.0 mg/l TDZ. Incorporation of 8 % coconut water in the above mentioned medium resulted satisfactory shoot growth and development. The best response towards root induction (90 %) was observed when 3-4 shoots clump were transferred onto halfstrength of gelled MS medium fortified with 4.0 mg/l IBA with 9.31 ± 1.45 roots per unit shoot. Successful acclimatization of well-rooted clumps of 4-5 shoots was achieved in a mixture of soil, sand, and compost (1 : 1 : 1) with 90% survival rate. Bangladesh J. Bot. 52(2): 337-343, 2023 (June)

Establishment of an Efficient Somatic Embryogenesis Protocol for Giant Reed (Arundo donax L.) and Multiplication of Obtained Shoots via Semi-Solid or Liquid Culture

This study developed an efficient protocol for the in vitro propagation of giant reed (Arundo donax L.) biomass, defining a complete cycle of the induction of somatic embryogenesis from immature inflorescences, followed by the maturation of somatic embryos and the subsequent multiplication of the derived shoots in liquid culture in a temporary immersion system (TIS). The best explants were found to be 30 cm long immature inflorescences, preferably collected in spring. Such an explant type was easy to decontaminate, and the spikelets isolated from it provided over 100 embryogenic callus lines. Among the callus induction media tested, gelled MS medium supplemented with 1.1 mg/L 2,4-D provided the highest percentage of responsive spikelets and the highest density of embryogenic callus. Maturation of the embryogenic callus was easily triggered on gelled MS medium devoid of plant growth regulators. The obtained shoots could be further multiplied on previously optimized gelled DKW medium supplemented with 30 g/L sucrose, 5 mg/L BA, 0.1 mg/L IBA, and 6.8 g/L plant agar. Subsequent high multiplication of the developed shoots was achieved in liquid culture in TIS using a Plantform™ bioreactor, with an immersion cycle of 12 min every 8 h.

In Vitro Propagation of Two Tomato Hybrids (Lycopersicon Esculentum Mill.) Via Tissue Culture Technique

This investigation was carried out at the Tissue Culture Lab., Biotechnology Research Center (BTRC) in Tripoli, Libya in 2009 to establish a micro propagation protocol for two hybrids of Tomato in which several factors were studied including the effect of concentration of growth regulator on shoot multiplication, and rooting of explant on MS media. Shoot tips were cultured into (MS medium) with different concentrations of benzyl adenine (BA) + 0.5 mg/l (IAA) compared with MS basal medium as a control treatment. Generally, the results showed that there were no significant differences between several concentrations of (BA+IAA) on the number of leaves and shoot length. The obtained results illustrated was M1 gave high number of shoots/explant, shoot length, and leaves number for two hybrids of Tomato which were 1.90, 3.89, 4.48 respectively compared with the control. The results showed that there were no significant differences between Tomato hybrids on shoot length and the number of shoots. The best result for leaves number was hybrid HH-56 which gave 4.54 leaves. From the foregoing results, MS media supplementing IAA at 0.2 mg/l was the best treatment for rooting stage of hybrid HH-56 and Hybrid AZHAR F1. The plantlets were transferred to plastic cups containing peatmoss + sand at 1:1 (V/V). Acclimatization of plantlets were successfully.

Validating the Efficacy of an Established Micropropagation Protocol for Commercial Propagation of Neolamarckia cadamba

Validating the Efficacy of an Established Micropropagation Protocol for Commercial Propagation of Neolamarckia cadamba

A Unified and Efficient Coordinating Framework for Autonomous DBMS Tuning

Recently using machine learning (ML) based techniques to optimize the performance of modern database management systems (DBMSs) has attracted intensive interest from both industry and academia. With an objective to tune a specific component of a DBMS (e.g., index selection, knobs tuning), the ML-based tuning agents have shown to be able to find better configurations than experienced database administrators (DBAs). However, one critical yet challenging question remains unexplored -- how to make those ML-based tuning agents work collaboratively. Existing methods do not consider the dependencies among the multiple agents, and the model used by each agent only studies the effect of changing the configurations in a single component. To tune different components for DBMS, a coordinating mechanism is needed to make the multiple agents be cognizant of each other. Also, we need to decide how to allocate the limited tuning budget (e.g., time and resources) among the agents to maximize the performance. Such a decision is difficult to make since the distribution of the reward (i.e., performance improvement) corresponding to each agent is unknown and non-stationary. In this paper, we study the above question and present a unified coordinating framework to efficiently utilize existing ML-based agents. First, we propose a message propagation protocol that specifies the collaboration behaviors for agents and encapsulates the global tuning messages in each agent's model. Second, we combine Thompson Sampling, a well-studied reinforcement learning algorithm with a memory buffer so that our framework can allocate the tuning budget judiciously in a non-stationary environment. Our framework defines the interfaces adapted to a broad class of ML-based tuning agents, yet simple enough for integration with existing implementations and future extensions. Based on extensive evaluations, we show that this framework can effectively utilize different ML-based agents and find better configurations with 1.4~14.1x speedups on the workload execution time compared with baselines.

Tissue culture of gerunggang (Cratoxylum arborescens (Vahl) Blume): multipurpose native species of Indonesian peatland

Gerunggang (Cratoxylum arborescens (Vahl) Blume) is a tree native to Indonesian peatlands that has great potential as an alternative fiber-producing species for the pulp and paper industry because it is adaptable and does not have the potential to disrupt local ecosystems. The species also contains important anti-cancer compounds for the pharmaceutical industry. These industries require large quantities of raw materials to meet their needs, which cannot be met by the supply of conventional plant seedlings. Tissue culture is one of the propagation biotechnologies that can be used as an alternative to provide more efficient germplasm. In this study, we developed a protocol for gerunggang propagation using tissue culture technique. The results showed that the combination of 1 mg/l 6-benzylamino purine (BAP) and 0.5 mg/l naphthalene acetic acid (NAA) in Murashige & Skoog media provided the fastest bud break time (2 weeks before the first subculture), with the highest shoot initiation (3.0 ± 0.2 cm), and after 4 weeks of incubation, the highest auxiliary shoot elongation (7.4 ± 0.1 cm). The significant effect of this hormone concentration also increases the nodule multiplication coefficient (NMC), which is 7.5. In vitro root response on the same propagation medium showed good growth with an average root length of 4.8 ± 0.2 cm. Acclimatization of plantlets was successful under ex vitro conditions and resulted in good growth after 2 and 3 months in the greenhouse. The advantage of applying the tissue culture method to gerunggang is that it can produce about seven times more seedlings from one explant in about 4 weeks. These results will be very useful for further research in the field and require a more intensive and comprehensive study.

Developing Paraphalaenopsis labukensis (Shim, A. Lamb & C.L. Chan), an Orchid Endemic to Sabah, Borneo, Asymbiotic Seed Germination and In Vitro Seedling Development

Paraphalaenopsis labukensis Shim, A. Lamb & C.L. Chan (P. labukensis) is an endangered monopodial epiphytic orchid threatened due to habitat fragmentation and overharvesting. Consequently, this research aimed to achieve in vitro propagation of P. labukensis through asymbiotic seed germination and seedling development. This study focused on identification and optimizing capsule maturity, basal media and nutrient requirements. In addition, after hand pollinating the plant, their capsules’ flowers, length, girth, and colour variations were recorded weekly to ascertain morphological maturity characters of this species. Murashige and Skoog (MS), Knudson C (KC), and Vaccin and Went (VW) basal media were used to determine seed vegetation at 60, 90, and 120 days. Subsequently, for the proliferation and development of protocorms, the best basal media were those supplemented with different organic additives (coconut water (CW) and banana homogenate (BH)) and plant growth regulators (PGRs). As a result, 120 days after pollination (DAP) was chosen as the best capsule age for germination since the capsules had reached maturity. The highest germination rate was attained after 90 days of culture in Knudson C (KC) medium, with 98.78 ± 0.89%, followed by Murashige and Skoog (MS) medium, with 92.80 ± 3.26%. The KC supplemented with 0.5 mg/L NAA media documented a maximum percentage of 17.25 ± 0.96%. The 5 g/L (w/v) BH recorded a higher protocorm proliferation percentage than CW. Additionally, after 150 days of culture (DOC), this medium combination produced the most leaves, averaging four or five. The result of this present study has successfully established an effective in vitro propagation protocol for this species.

The Root Collar Diameter Growth Reveals a Strong Relationship with the Height Growth of Juvenile Scots Pine Trees from Seeds Differentiated by Spectrometric Feature

This study is intended for forest owners considering options to increase the efficiency of the production of forest seedlings in automated nurseries. In the short rotation technology of the Scots pine (Pinus sylvestris L.), the production of seedlings was integrated in the process of grading seeds by spectrometric features, followed by the fall outplanting of containerized seedlings to a restorable post-pyrogenic site. There are studies of the spectrometric effect grading viable seeds, but a negligible amount of testing has been conducted on the seedling’s field performance from these seeds. It was very important for us to evaluate the morphogenesis of juvenile Scots pine trees in relation to the change in morphological features—increase in root collar diameter (RCD-increment) and increase in stem height (SH-increment)—in the second and third growing season from the field outplanting. To determine the nature of the interaction between independent (RCD increase), dependent (SH-increment), and categorical (seed spectrometric categories and timespans) variables, a regression analysis was performed using the R statistical software. All coefficients of the linear models are statistically significant at the 1% significance level, and all are positively associated with an increase in seedling height. On average, a RCD increment by 0.1 cm causes a SH-increment by 0.72 ± 0.16 cm (p = 9.779 × 10−6). In addition, in 2020, compared to 2019, the SH-increment was 5.46 ± 0.37 cm (p < 0.001). The seeds’ differentiation into spectrometric categories is strongly correlated with the stem height increment (p < 0.001). Thus, in order to improve the plant propagation protocol, it seems advisable to first condition the seeds in accordance with the spectrometric feature, and to then apply a coating (by pelleting or encapsulating) in order to improve the storage and seeding conditions. For a medium-term assessment of the short rotation technology with an integrated seed spectrometric grading in the future, the morphogenesis of Scots pine trees at 5, 7 and 10 growing seasons from the seedlings field outplanting is controlled.

Development of an Efficient and Rapid Micropropagation Protocol for In Vitro Multiplication of Maerua crassifolia Forssk

The difficult propagation of shrub and tree species and their extensive exposure to grazing threaten their abundance and lead to the necessity to find alternative means of propagation for these species. In vitro micropropagation techniques, viz., tissue culture, offer a promising tool for the rapid, cost-effective, and efficient propagation of different plant species. In the current study, a rapid and efficient in vitro multiplication protocol was developed for the micropropagation of Maerua crassifolia Forssk. Our results revealed that Murashige and Skoog (MS) medium with 7.5 µM of 6-benzylaminopurine (BA) and 1.0 µM of 1-naphthaleneacetic acid (NAA) led to the highest shoot formation (13.9 shoots per explant in 85.7% of the cultivated hypocotyls) among all other treatments. The best in vitro root formation was obtained on half-strength MS medium with 1.0 µM of indole 3-butyric acid (IBA) as 94.1% of the cultivated shoots formed 6.8 roots per microshoot on average. Ninety percent of the rooted plantlets were successfully acclimatized and are currently growing in the botanical garden of the Botany and Microbiology Department, King Saud University, Riyadh, Saudi Arabia. The genetic fidelity of the micropropagated plants was authenticated via flow cytometry. The results of the current study explained a simple, cost-effective, and efficient protocol for the micropropagation of the endangered M. crassifolia trees.

Control of media browning during micropropagation and assessment of biochemical and clonal fidelity of in vitro-derived and mother plants in Thottea siliquosa (Lamk.) Ding Hou., an important ethnomedicinal shrub

BackgroundThottea siliquosa (Lamk.) Ding Hou., an important medicinal shrub, is widely used in both ayurvedic and indigenous systems of medicine. Root being the most useful part, the plant is constantly uprooted and thus puts pressure on the natural population. Until date, no micropropagation study is available in this plant. The objective of the study is to develop an efficient in vitro propagation protocol and assessment of clonal fidelity of T. siliquosa. ResultsMedia browning was a serious issue during micropropagation, and the addition of 40.0 mg/L ascorbic acid reduced the media browning. For direct shoot regeneration, the optimum response (92% frequency with 20.9 shoots per explant) was obtained when 7-day-old cotyledons were cultured on WPM supplemented with 1.0 mg/L thidiazuron and 0.25 mg/L α-naphthalene acetic acid. The cultures were transferred to WPM augmented with 0.4 mg/L thidiazuron for shoot elongation and growth. On this medium, 100% of cultures responded with a mean number of 27.6 shoots. For callus induction, MS medium with 1.0 mg/L 2,4-dichlorophenoxyacetic acid and 0.5 mg/L N6-benzylaminopurin was used. Shoot organogenesis was initiated on the same medium, and calli with minute shoots were transferred to MS medium fortified with 0.5 mg/L N6-benzylaminopurin and 0.25 mg/L α-naphthalene acetic acid for highest shoot regeneration (100% cultures responded with a mean number of 26.5 shoots per explant). Maximum rooting frequency (82%) and number (20.8) were obtained on half-strength MS medium with 1.0 mg/L indole-3-butyric acid. The rooted plants were acclimatized and transferred to the field. The HPTLC and SCoT analysis revealed the phytochemical and clonal similarity between the in vitro propagated plants and mother plant. ConclusionsIn this study, it is confirmed that cotyledon is an excellent explant for direct and indirect shoot organogenesis in T. siliquosa. For direct shoot induction WPM and indirect organogenesis, MS medium was found to give better response. The true-to-type nature of in vitro-derived plants were confirmed by phytochemical and SCoT analysis. The protocol described here could be used for the large-scale propagation of elite clones of T. siliquosa.

Application of Cold Storage and Short In Vitro Germination for Somatic Embryos of Pinus radiata and P. sylvestris.

Somatic embryogenesis (SE) is an advanced vegetative propagation technology that, when used in combination with breeding and cryopreservation, offers the forest industry a powerful tool for the deployment of elite genotypes. Germination and acclimatization are critical and cost-intensive phases in the production of somatic plants. The efficient conversion of somatic embryos into robust plants is a necessity if a propagation protocol is to be successfully adopted by the industry. In this work, these late phases of the SE protocol of two pine species were investigated. A shortened germination protocol and more controlled acclimatization were investigated for Pinus radiata, testing embryos from 18 embryogenic cell lines. A more simplified protocol, including a cold storage phase, was also compared among 10 of these cell lines. A shortened germination period and more controlled protocols significantly improved the acclimatization of somatic embryos directly from the lab to the glasshouse. When results for all cell lines were pooled, there were significant improvements in all growth characteristics (shoot height, root length, root collar diameter, and root quadrant score). When the more simplified protocol involving cold storage was tested, improvements were seen in the root architecture. For Pinus sylvestris, the late phases of somatic embryogenesis were investigated on seven cell lines in a set of two trials (four to seven cell lines per trial). During the germination phase, a shortened and simplified in vitro period, a cold storage option and basal media were explored. Viable plants were obtained from all treatments. However, there is still the need to improve germination and related protocols together with growing regimes for Pinus sylvestris. The improvements to protocols presented here, particularly for Pinus radiata, result in greater survival and quality of somatic emblings, leading to reduced costs and increased confidence in the technology. Simplified protocols using a cold storage option show great promise and, with some further research, could lead to reductions in the cost of the technology.

Quality Evaluation of Mustard Microgreens Grown on Peat and Jute Substrate

Consumers appreciate microgreens for their vast variety of colors and flavors. Usually, they are grown employing peat, a substrate that is used in large quantities. In order to identify a more sustainable propagation protocol and to reduce the amount of peat consumed, alternative propagation protocols were evaluated. Jute is a biodegradable substrate with lower post-harvest costs because it does not leave particles on microgreens. This work evaluates the microgreen yield, flavor, texture, and phytochemical compounds when grown on jute. Green mustard (Brassica nigra) is one of the most popular microgreens. When growing these microgreens on jute (three repetitions), it was necessary to increase the frequency of irrigation and reduce the amount of water for each turn. In addition, the propagation time needed to be increased from 5 to 7 days. The tasters found no difference in flavor and only a slight difference in texture was observed when microgreens were grown on jute. The phenol and chlorophyll levels were unchanged, while carotenoid levels were slightly higher. Thus, the cultivation of green mustard on jute has a minimal impact on microgreens and leads to increased sustainability and reduced post-harvest costs.

Comparative assessment of morphological, physiological and phytochemical attributes of cultivated Valeriana jatamansi Jones in Uttarakhand, West Himalaya

Medicinal plants are global sources of herbal products, drugs, and cosmetics. They are disappearing rapidly due to anthropogenic pressure, overexploitation, unsustainable harvesting, lack of knowledge on cultivation, and the availability of quality plating materials. In this context, standardized in-vitro propagation protocol was followed to produce Valeriana jatamansi Jones, and transferred in two locations at Kosi-Katarmal (GBP) Almora (1200 masl) and Sri Narayan Ashram (SNA) Pithoragarh (Altitude 2750 masl), Uttarakhand. Over the three years of growth, plants were gathered from both locations for determining biochemical and physiological parameters, and growth performance. The plants growing at Sri Narayan Ashram (SNA) showed considerably (p < 0.05) higher amounts of polyphenolics, antioxidant activities, and phenolic compounds. Similarly, physiological parameters (transpiration 0.004 mol m−2 s−1; photosynthesis 8.20 μmol m−2 s−1; stomatal conductance 0.24 mol m−2 s−1), plant growth performance (leaves number 40, roots number 30, root length 14 cm) and soil attributes (total nitrogen 9.30; potassium 0.025; phosphorus 0.34 mg/g, respectively) were found best in the SNA as compared to GBP. In addition, moderate polar solvent (i.e., acetonitrile and methanol) was found suitable for extracting higher bioactive constituents from plants. The findings from this study revealed that large-scale cultivation of V. jatamansi should promote at higher elevation areas such as Sri Narayan Ashram to harness the maximum potential of the species. Such a protective approach with the right interventions will be helpful to provide livelihood security to the local populace along with quality material for commercial cultivation. This can fulfill the demand through regular supply of raw material to the industries and simultaneously promote their conservation.

In vitro Propagation of Endemic Species Mahonia Jaunsarensis Ahrendt Through Callus Culture.

Mahonia jaunsarensis Ahrendt (Family Berberidaceae) an endemic species was successfully propagated in vitro. An efficient propagation protocol has been developed first time. The callus cultures were established from leaf explants on Murashige and Skoog (MS) medium supplemented with 2,4-Dichlorophenoxyacetic acid (2,4-D; 1µM) and resulted 70% callus induction with green compact callus. When callus was transferred to MS medium containing Thidiazuron (TDZ; 0.75µM), maximum average number of shoot (3.06) produced but shoot length (3.37cm) and average leaf number (2.87) was increased upon transfer to MS medium containing N6-benzylaminopurine (BA; 6.0µM) plus α-naphthalene acetic acid (NAA; 0.5µM). In MS medium containing indole-3-butyric acid (IBA; 0.01µM), the maximum rooting percentage (56%) and average root number (2.56) per shoot and root length (3.33cm) were recorded. The rooted plantlets transferred in vermiculite + garden soil + farmyard manure (1:1:1) with maximum (55%) survival percentage under greenhouse condition. The phytochemical analysis of leaves obtained from tissue culture-raised plants revealed significantly higher levels of alkaloids (berberine and palmatine) than those obtained from wild plants. Similar trends were observed for antioxidant and antimutagenic activities. Results of this study offer a baseline for the conservation and sustainable utilization strategies forM. jaunsarensis.

Rapid and Efficient In Vitro Propagation Protocol of Endangered Wild Prickly Pear Growing in Eastern Morocco

The Opuntia genus is widely recognized as a significant member of the Cactaceae family. The eastern Moroccan prickly pear’s wild ecotype is renowned for its production of fruits of superior quality, which are in high demand. Nonetheless, the white cochineal (Dactylopius opuntiae) has emerged as a significant hazard to the persistence of the indigenous wild prickly pear population in the majority of the country’s territories. Conventional plant propagation techniques may facilitate the transmission of pathogens to successive generations and thus fail to satisfy market requirements. Therefore, the primary goal of this study was to develop a rapid and efficient protocol for large-scale production of the eastern Moroccan wild ecotype using axillary buds as the starting material. Sterilization of the starting material is a crucial but challenging step in this species, as the meristem is located just beneath the spine. The protocol developed for this study produced moderately satisfactory results, with 20 to 30% contamination in each experiment. The obtained shoots were incubated on Murashige and Skoog medium supplemented with varying concentrations of BAP (0, 1.5, 3, 5, and 7.5 mg/L). The treatment with BAP at 5 mg/L exhibited a statistically significant increase in the average number of regenerated shoots per explant (19.42). The effect of kinetin on the rooting of prickly pear was evaluated by transferring the shoots to a MS medium supplemented with varying concentrations of kinetin (0, 0.5, 1, 1.5, 2, and 2.5 mg/L). The use of kinetin increased the number and length of roots while also shortening the root development period from 21 days to 10 days. The best results were obtained at a concentration of 1.5 mg/L of kinetin. Furthermore, satisfactory acclimatization of plants was achieved by using plastic containers with a gradually increasing opening of the lids. The outcomes of this experimentation have significant potential to facilitate the preservation of this botanical variety, reduce the risk of white cochineal infestation, and address the need for superior quality fruit supply in the market.

Establishment of a suitable regeneration protocol for rapid propagation of Piper nigrum L. through in vitro culture

An experiment was performed to establish a suitable regeneration protocol for the rapid propagation of Piper nigrum L. using nodal segments and leaf tissues as explants through direct and indirect organogenesis. After surface sterilization, several types of explant were inoculated onto gelled MS medium containing various concentrations and combinations of growth regulators for callus and direct shoot induction. The highest callus induction frequency was 92% and 84% in the case of leaf tissues and nodal segments, respectively, in gelled MS medium containing 2.0 mg/l 2,4-D. Multiple shoots (6.43±0.35 shoots per unit callus) were obtained when the calli from both explant types were cultured on MS medium containing 1.0 mg/l BAP. Nodal segments showed the best result (85%) in terms of direct shoot induction in MS medium supplemented with 1.5 mg/l BAP, where the highest number of shoots per explant was 5.03±0.69. The directly induced shoots were multiplied and elongated on MS medium containing 1.0 mg/l BAP and 0.5 mg/l IBA, the number and length of regenerated shoots per culture being 6.07±0.39 and 5.84±0.65 cm, respectively. The best response to root induction (86.67%) was observed when shoots were transferred to ½ strength of gelled MS medium supplemented with 1.5 mg/l IBA within 16-24 days, with 13.60±1.76 roots per shoot unit. The well-rooted shoots were successfully acclimated in a mixture of soil, sand, and compost (1:1:1) with a survival rate of 88%.&#x0D; Jahangirnagar University J. Biol. Sci. 11(1 &amp; 2): 31-40, 2022 (June &amp; December)

A Complete Micropropagation Protocol for Black-Leaved Zamioculcas zamiifolia (Lodd.) Engl. ‘Dowon’

Zamioculcas zamiifolia, a drought-resistant plant in the family Araceae, is a popular ornamental potted foliage plant originating from tropical east and subtropical southeast Africa. The growth and propagation rate of this species is low in conventional propagation methods. Therefore, the current study aimed at developing a complete in vitro propagation protocol of black-leaved Raven® ZZ plant (Z. zamiifolia ‘Dowon’)—a novelty on the floricultural market. In order to initiate an axenic culture, the disinfection of leaf explants was performed with sodium hypochlorite and mercury chloride. Next, leaf segments were cultured on the Murashige and Skoog (MS) medium with the addition of 6-benzyladenine (BA) and/or ɑ-naphthalene acetic acid (NAA) at various concentrations. The highest number of shoots (11) and leaves (22) per explant was obtained in a medium enriched with 2 mg·L−1 BA together with 0.5 mg·L−1 NAA. The maximum number of roots (3.33) was produced in microshoots cultured on the medium supplemented with 2 mg·L−1 NAA. On the other hand, the longest roots (2.66 cm) were produced on a medium containing 2 mg·L−1 NAA and 0.5 mg·L−1 BA. The combination of 0.5 mg·L−1 BA and 0.5 mg·L−1 NAA was most effective in stimulating callus formation (78.33%). Rooted plantlets were transferred to plastic pots filled with coco peat and acclimatized to ambient greenhouse conditions with an average 68.19% survival rate. This is the first report on a complete micropropagation protocol of black-leaved zamioculcas.

Propagation of Medicinal Plants for Sustainable Livelihoods, Economic Development, and Biodiversity Conservation in South Africa.

South Africa is blessed with vast plant resources and unique vegetation types. Indigenous South African medicinal plants have been well-harnessed to generate income in rural communities. Many of these plants have been processed into natural products to heal a variety of diseases, making them valuable export commodities. South Africa has one of the most effective bio-conservation policies in Africa, which has protected the South African indigenous medicinal vegetation. However, there is a strong link between government policies for biodiversity conservation, the propagation of medicinal plants as a source of livelihood, and the development of propagation techniques by research scientists. Tertiary institutions nationwide have played a crucial role in the development of effective propagation protocols for valuable South African medicinal plants. The government-restricted harvest policies have also helped to nudge natural product companies and medicinal plant marketers to embrace the cultivated plants for their medicinal uses, and thus have helped support the South African economy and biodiversity conservation. Propagation methods used for the cultivation of the relevant medicinal plants vary according to plant family and vegetation type, among others. Plants from the Cape areas, such as the Karoo, are often resuscitated after bushfires, and propagation protocols mimicking these events have been established through seed propagation protocols with controlled temperatures and other conditions, to establish seedlings of such plants. Thus, this review highlights the role of the propagation of highly utilized and traded medicinal plants in the South African traditional medicinal system. Some valuable medicinal plants that sustain livelihoods and are highly sought-after as export raw materials are discussed. The effect of South African bio-conservation registration on the propagation of these plants and the roles of the communities and other stakeholders in the development of propagation protocols for highly utilized and endangered medicinal plants are also covered. The role of various propagation methods on the bioactive compounds' composition of medicinal plants and issues of quality assurance are addressed. The available literature, media online news, newspapers, and other resources, such as published books and manuals, were scrutinized for information.