Combinations Of NAA Research Articles

Effects of plant growth regulators on the callus of rose leaves

In the present study effects of different plant growth regulator combinations on leaf callus growth in two Rosa hybrida cultivars, Rosa hybrida ‘Ingrid Bergman’ and Rosa hybrida ‘Xindongfang’, were analyzed. The plant growth regulators were added into MS, MS + 2,4-D (1.5 mg/l, 2.5 mg/l, 3.5 mg/L, and 4.5 mg/l) and MS + 2.0 mg/l NAA + 1.0 mg/l ZT, MS + 2.0 mg/l NAA + 1.0 mg/l KT, MS + 2.0 mg/l NAA + 1.0 mg/l 6-BA and MS + 2.0 mg/l NAA. The leaves were treated as broken and cultured on these nine different media types. Results showed significant differences in color, morphology, and callus quantity. In the first medium, the callus of rose leaves was the best on MS + 4.5 mg/l 2,4-D medium, followed by MS + 2.5 mg/l 2,4-D and MS + 3.5 mg/l 2,4-D, and finally MS + 1.5 mg/l 2,4-D. No callus was found in MS medium. In the second kind of medium, the leaves grow callus, and the callus grew best on MS + 2.0 mg/l NAA + 1.0 mg/l ZT medium, followed by MS + 2.0 mg/l NAA + 1.0 mg/l KT, finally MS + 2.0 mg/l NAA + 1.0 mg/l 6-BA, MS + 2.0 mg/l NAA medium, the callus grew loose and dry, and lost luster after a few days. Therefore, 2,4-D, KT, ZT, 6-BA, and NAA can promote the formation of callus. With the increase of concentration of 2,4-D, the callus induction rate increased, and the color of the callus also changed significantly. The combination of KT, ZT, and NAA had an obvious effect on callus induction and expansion. Bangladesh J. Bot. 52(3): 867-873, 2023 (September)

Optimization of in vitro propagation of jamun variety Konkan Bahadoli

Micro-propagation is a vital technique for round-the-year clonal multiplication of plants. Jamun is anindigenous fruit tree with great pharmacological importance. Two explants viz., shoot tips and nodal explants were cultured from January to May. The shoot regeneration from nodal explants was tested using 6-benzyl amino purine (BAP; 1.0-4.0 mg l-1) and α-naphthalene acetic acid (NAA; 0-0.25 mg l-1), while rooting was investigated with NAA (0.5-2.0 mg l-1). Maximum explant establishment was recorded in May. The nodal explants showed better establishment response (50.0%) than shoot tips (40.7%). The maximum shoot regeneration (72.16%) and earliest evocation response from nodal segments (14.0 days) were obtained with combinations of BAP (2.0 mg l-1) and NAA (0.25 mg l-1). Silver nitrate (AgNO3 @ 3.0 mg l-1) enabled shoot proliferation. Maximum rooting occurred on the medium containing 0.5 mg l-1 NAA. The in vitro plants were hardened on the potting mixture of cocopeat (4): perlite (1): vermiculite (1).

Novel System for the Production of the Bioactive N-alkylamide ‘Spilanthol’ Through Somatic Embryogenesis in Acmella ciliata Kunth (Cass.)

Acmella ciliata Kunth (Cass.), a medicinally important plant in the family Asteraceae, has high commercial value because of its traditional phytomedicinal uses. The plant contains many phytochemicals like alkyl amides, alkaloids, tannins, saponins and flavonoids accountable for most of its pharmacological applications. The study presented here reports the callus culture and somatic embryogenesis of this plant thereby raising a novel system for the subsequent production of the N-alkyl amide ‘spilanthol’, the valuable secondary metabolite presents in it. Murashige and Skoog (MS) medium supplemented with auxins either alone or in combination with cytokinins were used for the induction and maturation of somatic embryos. MS medium supplemented with 2,4-D (0.5, 1.0 and 2.0 mg.L-1) produced black friable callus whereas, 1.0 mg.L-1 NAA in combination with 0.5 mg.L-1 BA induced white, slightly purple coloured friable callus which on further subculture to fresh medium induced somatic embryos that germinated into plantlets upon transfer to MS basal medium. The mode of regeneration via somatic embryogenesis was confirmed by histological analysis through free-hand sectioning and stereomicroscopic observation. The plantlets raised through somatic embryogenesis after a short hardening period, were found to acclimatise in the field at 83.33% efficiency and exhibited genetic uniformity with 96.6% similarity in the ISSR analysis. HPLC analysis of in vitro raised embryogenic callus showed 239.512 µg.g-1 spilanthol content which was comparatively higher than the mother plants (92.19 µg.g-1). The bioproduction of the N-alkylamide ‘spilanthol’ through embryogenic callus can be extended for the scale-up production of this bioactive compound using bioreactor technology for the formulation of phytodrugs.

A prospective insight into the reproduction of Sternbergia clusiana K. Gawl.: an endangered species

Sternbergia clusiana (SC) is a bulbous plant having an alluring ornamental flower, medicinal properties, and an endangered status due to climate-change ecology. However, the methods of propagation for conserving this notable species have been neglected. The main aim of this study was to evaluate the influence of plant growth regulators (PGRs) on SC regeneration by virtue of a asexual propagation method, with the intention of preserving this endangered species from extinction. In this regard, having collected the bulbs from a wild habitat, bulbs were cut in accordance with the chipping technique. Thereafter, pieces were treated with 6-benzylaminopurine (BAP) at 250, 500, and 1000 mg·L−1, 1-naphthaleneacetic acid (NAA) at 100, 250, and 500 mg·L−1, putrescine (PU) at 50, 100, and 200 mg·L−1, BAP and NAA combination, BAP and PU combination, and a control. Pieces were then transferred into primary and terminal growing media under laboratory and greenhouse conditions, respectively. Next, the number, length, width, and weight of bulbs were determined. The greatest bulb number (2.10 counts), bulb length (26.75 mm), bulb width (16.25 mm), and bulb weight (3.13 g) were observed in the PU treatments. In addition, the results showed that even though the treated bulbs with BAP and PU combinations had preferable performance generally, the results were inconsistent. Taken together, this research indicates that utilizing asexual propagation along with PGRs, in the proper dosage, can serve as a mechanism to protect the endangered geophyte from extinction in nature.

Effect of Different Hormones Concentration on In vitro Regeneration of Apricot Cultivars

The use of modern breeding methods along with appropriate cultivation techniques facilitates the acquisition and multiplication of varieties that possess the desired characteristics. Therefore, efforts towards the in vitro propagation of woody plants are increasing day by day. Today, plants such as Malus, Prunus, Pyrus, Ribes, Rubus, etc., can be successfully propagated in vitro. Apricot stands out as a stubborn species among Prunus types for shoot regeneration and genetic transformation. In this context, this study aims to determine how different plant growth regulators affect shoot regeneration of some native apricot varieties, which hold significant importance in apricot cultivation in our country. In the conducted study, mature cotyledons of Kabaaşı, Hacıhaliloğlu, and Hasanbey apricot varieties were used along with the culture medium consisting of MS. Different doses and ratios of plant growth regulators, including BAP and TDZ, in combination with NAA and GA3, were added to the culture media. At the end of the in vitro study, the callus and primordium formation rate (%), bud and shoot formation rate (%) and number of shoots per explant were recorded. According to the results, the variety with the highest callus formation was Kabaaşı, followed by Hasanbey and Hacıhaliloğlu. In all three varieties, the rate of callus formation decreased in media containing GA3. Regarding the stage of shoot regeneration from callus, the highest shoot formation with an average of 4 shoots per explant was observed in the Kabaaşı variety in the TDZ (1.0 mgL-1) + NAA (0.25 mgL-1) and TDZ (1.0 mgL-1) + NAA (0.50 mgL-1) media. Looking at the other varieties, the highest number of shoots, 1.6 shoots per explant, was obtained from the TDZ (2.0 mgL-1) + NAA (0.25 mgL-1) medium in Hasanbey and Hacıhaliloğlu varieties. As a result of the findings, the Kabaaşı variety showed the best result in terms of the regeneration capacity of apricot varieties. In contrast the best regeneration medium was obtained from the combinations of TDZ and NAA.

Effect of Adding NAA and BAP on the Growth of Kepok Kuning Banana (Musa paradisiaca L.) Planlet on MS Media in Vitro

The availability of Kepok Kuning Banana (Musa paradisiaca L.) seeds is increasingly limited due to physiological diseases such as leaf wilt. Therefore, tissue culture techniques were carried out as a solution for propagating Kepok Kuning banana seeds which disease free. This study's purpose is to determine the effect of adding NAA and BAP to MS media in vitro on the growth of Kepok Kuning Banana plantlets. This study used a completely randomized design with one treatment factor, the combination of NAA and BAP concentrations with five treatments; A (4ppm NAA + 8ppm BAP), B (5ppm NAA + 7ppm BAP), C (6ppm NAA + 6ppm BAP), D (7ppm NAA + 5ppm BAP), E (8ppm NAA + 4ppm BAP). Plantlet growth data including number of shoots, leaves, roots, and plantlet height were analyzed using one-way ANOVA test followed by Duncan's test at 5% level. The results showed that the addition of NAA and BAP affected the growth of Kepok Kuning banana plantlets on MS media in vitro. This is shown in the optimal number of shoots and leaves in treatment A, treatment B, and treatment C. Meanwhile, the number of roots and plantlet height is optimal in treatment D, and treatment E.

The Effect of Immersing the Bud Cuttings with a Combination of NAA and BA Growth Regulators and the Addition of Optimus Plus Nanofertilizer in Improving some Vegetative Qualities of Ber Seedlings Apple Variety

The study was carried out during the growing season 2022 in one of the orchards located in Al-Sajla area / Al-Khairat district / Holy Karbala governorate On seed Sidr varieties two years old. To find out the effect of immersing bait pens, an apple cultivar was treated with a combination of growth regulators in three concentrations (0, 40 + 20, 40 + 80 mg.L-1) and symbol (A0, A1, A2) in sequence, and the addition of Optimus Pius nano fertilizer in three concentrations (0, 20, 40 m. L-1) and its symbol (B0, B1, B2) sequentially and the overlap between them in the vegetative traits of Sidr seedlings, apple cultivar. The experiment was carried out according to the randomized complete block design (RCBD) as a factorial experiment with three replications. The results showed that immersing the grafting pen with a combination of growth regulators at A1 concentration had a significant effect on vegetative traits (percentage of vaccination success, main stem length, main stem diameter, number of lateral branches, number of leaves and leaf area) amounted to (88.900%, 142.567 cm, 40.047 mm 17.447 branch. Sapling, 827 leaf. Sapling, 22.567 cm2. sapling-1) sequentially compared to the comparison treatment, Ground fertilization with nano-fertilizer at a concentration of B2 also led to a significant increase in (percentage of vaccination success, main stem length, main stem diameter, number of lateral branches, number of leaves and leaf area) amounting to (88.900%, 144.900 cm, 40.150 mm, 17.667 branch. Sapling, 828 leaves. Seedling-1, 22.967 cm2. sapling-1) sequentially compared with the comparison treatment. The bilateral interaction between the two study factors also led to a significant effect on all vegetative characteristics of Sidr seedlings of an apple variety, especially when treated (A1 B2) compared to the comparison treatment of the overlap.

In vitro propagation and genetic fidelity analysis of Celastrus paniculatus Willd – a vulnerable high trade medicinal woody liana of Western Ghats

Celastrus paniculatus Willd, a vulnerable important woody liana with high medicinal value and limited reproductive capacity has been over exploited due to its demand in the pharmaceutical industry. In the present study, an efficient micropropagation protocol was developed from nodal explants on Murashige and Skoog and woody plant medium. The effect of various strengths of WP and MS medium (total, half strength) and different concentrations of plant growth regulators including cytokinins [6-benzylaminopurine (BAP) and Kinetin) and auxins (α-naphthaleneacetic acid (NAA) and (IBA) indole-3-butyric acid] on proliferation and rooting was investigated. C. paniculatus shoots showed maximum proliferation in WP medium supplemented with BAP (0.5 mg l-1). Best rooting frequency and higher root number and length were attained in medium fortified with NAA in combination with 0.2% activated charcoal. Seventy percent of the plantlets were successfully acclimatized to ex vitro conditions, exhibiting normal development. These plantlets can be used to replenish the declining populations in the wild. Molecular profiling of the in vitro developed plantlets was done using RAPD to confirm their genetic uniformity.

Response of Different Safflower Genotypes to Anther Culture

In this study; the responses of 12 safflower genotypes to anther culture were determined in MS medium containing increasing doses of BAP (0, 0.5, 1, and 2 mg L-1) and NAA (0, 0.1, and 0.5 mg L-1). Donor plants were grown under field conditions and the main capitulas were collected in early or late uninucleate stage. A pre-treatment was applied to their capitules at +4 °C for 3-4 days. Only cvs. Balcı and Linas produced regenerable callus among these genotypes. Although regenerable callus rate was higher in Balcı than Linas, shoot regeneration was achieved only in Linas with 0.5 or 1 mg L-1 of BAP + 0.1 mg L-1 of NAA combinations. In total, 8 shoots of Linas were moved to ½ MS medium consisting of 0.1 mg L-1 of NAA, but rooting of them could not be performed. In sum, among the 12 safflower genotypes, cvs. Balcı and Linas have higher anther culture regeneration potential; however, it is necessary to focus on shoot development and rooting on these cultivars in further experiments.

Regeneration of safflower genotypes through callus mediated organogenesis using cotyledonary node explants

In this study, the cotyledon nodes of five safflower genotypes (Balcı, Linas, inbred lines 24, 25, and 55) were cultured for shoot regeneration via organogenesis in MS medium involving different TDZ (0.1, 0.5, and 1 mg L-1) and NAA (0, 0.2, and 0.5 mg L-1) doses. The highest rate of shoot forming calli was obtained from genotype 25 in all NAA and TDZ combinations, and there was no statistical difference between genotypes 24 and 25. The number of shoots per callus was found to be low in genotypes with a high rate of shoot forming calli. The maximum shoot number was obtained from the cultivar Linas on medium containing 1 mg L-1 TDZ, with 9.6 shoots/per callus and this value was followed by cultivar Balcı cultured at the same dose with 6.7 shoots/per callus. The rooting of safflower genotypes differed depending on the NAA content of the medium. Better rooting was achieved on medium with 2 mg L-1 NAA for Balcı, 1 mg L-1 NAA for Linas, and 0.1 mg L-1 NAA for genotypes 25 and 55. On the other hand, genotype 24 indicated rooting only on medium with 2 mg L-1 NAA, but it was very low. As a result; regeneration of safflower genotypes via callus-mediated organogenesis from cotyledonary explants was varied depending on TDZ and NAA doses, and many shoots were induced in Linas cultivar at 1 mg L-1 TDZ. However, the rooting of the regenerated shoots was quite low at different NAA doses.

Callus Induction and Regeneration Response of Mustard (Brassica spp.)

Mustard (Brassica spp.) is the third most important edible oil crop belonging to the family Brassicaceae (Cruciferae) and the genus Brassica. Due to the growing world population and increasing industrialization, the demand for edible oil and biofuels is increasing worldwide. Stress-tolerant mustard variety creation by transformation or genome editing procedures, as well as varietal in vitro regeneration responses, are critical challenges. The objective of the study was to design a tissue culture protocol for better use the explant by using hypocotyls and cotyledon explants to observe the in vitro response to callus induction and regeneration of three mustard types, namely BARI Sarisha-14, Binasarisha-4, and Binasarisha-9.Explants were cultured on Murashige and Skoog medium (MS) supplemented with different concentrations and combinations of NAA(10 mg mL-1), BAP (10 mg mL-1 for callus induction and shoot initiation medium , 2.5 mg mL-1 for shoot outgrowth medium), AgNO3(50 mg mL-1), GA3(2 mg mL-1), and IBA (10 mg mL-1)for callus induction, shoot initiation, shoot outgrowth, and root initiation. Binasarisha-4 showed the best performance with hypocotyls76.67% callus induction, 70.03% shoot initiation, 92.23% shoot outgrowth, and 40% root initiation followed by BARI Sarisha-14 (71.67%, 58.23%, 73.33% and 23.33% respectively) and Binasarisha-9 (54.17%, 38.87%, 45.83% and 11.11% respectively). It is recommended that in vitro hypocotyls culture of Binasarisha-4 can be used for frequent and successful genetic transformation in Mustard.

THE GROWTH OF PIMPINELLA ALPINA HOST CALLUS AT VARIOUS TREATMENTS OF PLANT GROWTH REGULATOR CONCENTRATIONS OF NAA. 2,4 D AND ITS COMBINATION WITH BAP

Purwaceng (Pimpinella alpina) is an herbal plant that has aphrodisiac, diuretic, and tonic properties. The imbalance between the growth and the exploitation of this plant makes it included in Appendix I (endangered), so its conservation will be crucial. One method of the conservation of this plant is tissue culture. This method is considered one way to induct the callus. The research objective is to find the effect of various treatments of plant growth regulator concentrations of NAA. 2,4 D and its combination with BAP on the growth of purwoceng callus. Concentration of plant growth regulator NAA (0.5 mg L-1, 1 mg L-1, 2 mg L-1), 2,4-D (0.4 mg L-1, 0.8 mg L-1, 1.2 mg L-1 ), a combination of NAA with BAP (0.5 mg L-1 NAA: 1 mg L-1 BAP, 1 mg L-1 NAA: 2 mg L-1 BAP, mg L-1 NAA: 3 mg L-1 BAP) and a combination of 2, 4- D with BAP (1 mg LL 2,4-D : 0.5 mg L BAP, 1 mg L-1 2,4-D : 1 mg L-1 BAP, 2 mg L-1 2,4-D : 1 mg L-1 BAP) into Murashige Skoog medium. Then purwaceng leaf explants were planted on the medium until the growth was seen. This research shows that the plant growth regulator concentration has a different influence on the growth of callus that is recognized by the day of callus induction, colour and callus texture. MS media with growth regulator 1.2 mg L-1 2,4-D can grow callus faster (17 days) with friable texture.

In Vitro Multiplication of Lophostemon suaveolens (Sol.ex Gaertn.) Peter G.Wilson & J.T. Waterh): Peatland Tree Species for Rehabilitation

Peatlands in Indonesia are one of the world’s largest carbon sinks, helping to regulate greenhouse gas emissions and global climate change. Lophostemon suaveolens is a relatively unexplored plant found in Papua’s endemic peat ecosystem that grows well in wet areas with low fertility. It is geographically dispersed and has the potential for peatland rehabilitation. Seed is one of materials for the reproduction of L. suaveolens. However, the difficulty in seed collection and the limitation in seed production has become a current problem for its cultivation. Seed multiplication by using an in vitro method would be one of the mechanisms to overcome the problem. We present an efficient and reproducible protocol for in vitro multiplication of plantlets using nodal segments and shoot apices collected from plantlets. After 3 months of the culture initiation stage, the elongated axillary shoots were separated from the clumps and further multiplied using Murashige and Skoog (MS) media supplemented with (1) BAP (0.5 mL/L) as single PGR, (2) NAA (0.1 mL/L) as a single PGR, and (3) a combination of two types of PGR BAP (0.5 mL/L) and NAA (0.1 mL/L). Up to an incubation period of 6 months, the efficiency of leaf axillary shoot propagation was determined by counting the number of nodule multiplication coefficient (NMC), shoot length, root length, and number of leaves (six consecutive subcultures). The higher the NMC, the higher the plantlets obtained, increasing shoot regeneration from nodules physiologically increasing evapotranspiration in vitro. The highest of NMC (8.4) was observed in MS medium with a combination of 0.5 mL/L BAP and 0.1 mL/L NAA (double PGRs), with the longest shoots (5.91 cm), the longest root length (8.83 cm), and the most leaves (32). When a combination of BAP and NAA were used simultaneously, the plantlets during acclimatization were the highest survived. It was concluded that MS in combination with 0.5 mL/L BAP and 0.1 mL/L NAA is the most appropriate protocol for the success of in vitro multiplication of L. suaveolens. This is the first report of L. suaveolens in vitro multiplication, and the protocol could be used to propagate this peatland species on a large scale. The authors acknowledge the limitations of the experimental work and recommend further work to increase the sample size and complete the field-testing phase to help verify the initial findings presented in this paper.

High‑frequency plant regeneration and genetic fidelity assessment of regenerants by molecular and biochemical markers in Woodland Sage (Salvia nemorosa L.)

Salvia nemorosa L. is a valuable medicinal and ornamental plant from Lamiaceae . A high-frequency protocol for in vitro regeneration from S. nemorosa leaf explants was developed based on our results. Various plant growth regulators were evaluated for their effects on direct and indirect organogenesis , callogenesis, shoot multiplication, and rooting of explants at different concentrations. The maximum percentage of direct shoot regeneration (97.5%) and the number of shoots per explant (8.6) were obtained on MS medium supplement with 8 µM TDZ plus 8 µM NAA . The result revealed that media amended by 16 µM 2, 4-D with 8 or 16 µM BA was the best treatment for callus induction. Media containing the combination of 8 µM TDZ plus 2 µM NAA provided the highest rate of shoot induction (84%) and the highest number of shoots (5.4) from calli. Regenerated shoots were multiplied efficiently with an average of 7.4 shoots and 32.4 leaves per explant on media containing 12 µM TDZ. In the present study, TDZ and BA treatments led to the highest proliferation rate; however, adenine sulfate was preferred for proliferation because it did not produce hyperhydric plants. The genetic fidelity of 19 plantlets (one donor plant, nine direct and nine indirect regenerants) was appraised by 10 ISSR markers. The molecular analysis revealed that direct organogenesis produced true-to-type plantlets and could be employed for clonal multiplication of Woodland Sage. Indirect regenerants showed polymorphic bands (12.5–33.33%), confirming the somaclonal variation occurrence during indirect organogenesis. Also, HPLC analysis of regenerants showed that two indirect regenerants accumulated higher rosmarinic acid content than mother plants, while three of them accumulated lower rosmarinic acid content. As part of this study, molecular and biochemical markers have been employed to investigate somaclonal variation in in vitro regeneration of the Salvia genus for the first time, which could be utilized to develop reliable methods for multiplying elite lines and achieving high-yield lines. • A high-frequency protocol for S. nemorosa in vitro regeneration was developed. • TDZ and NAA combination produced plantlets with high genetic and biochemical fidelity. • Adenine Sulphate prevents the Hyperhydricity in regenerated plantlets. • The indirect regeneration affected PAL activity, rosmarinic acid, and genetic content.

Elimination of coexisting canna yellow mottle virus, bean yellow mosaic virus and cucumber mosaic virus from Canna generalis cv. black knight through in vitro chemotherapy of rhizome explants.

During our previous study, the mixed infection of canna yellow mottle virus (CaYMV), bean yellow mosaic virus (BYMV), and cucumber mosaic virus (CMV) was identified in a Black Knight cultivar of canna exhibiting severe yellow streak and mottling symptoms. Before the development of the virus-free plants, the ability of callogenesis and organogenesis from the ovary, stalk, and rhizome explants was tested on different concentrations and combinations of TDZ, NAA, BAP, and Ads growth regulators. The performance of rhizome explants was above all the explant types and 33.33 ± 1.67 rhizomes (out of 50 placed) showed callus development on ME medium (MS supplemented with 0.8mg/L TDZ and 0.25mg/L NAA) and further on a refined M4 medium (MS supplemented with 4.0mg/L BAP, 1.0mg/L NAA and 50mg/L Ads) produced 4.06 ± 0.16 shoots per explant. The development of virus-free plants was attempted by in vitro chemotherapy using ribavirin. Not only in callogenesis and shoot development but also in the ribavirin treatments, rhizomes developed about 3.78 ± 0.68 shoots per explant on 40mg/L ribavirin in the ME medium. These optimizations suggested that ME medium for callogenesis, M4 medium for shoot development and the treatment of 40mg/L ribavirin for 30days at M4 medium was effective. The elimination of coinfection of all three viruses from rhizome explants of 0.5 cm2 of the Black Knight cultivar was attempted. Consequently, a total of 53.33% of plants free from all three viruses (48 out of the 90 plants developed) were obtained when screened by RT-PCR and PCR for their absence.

In vitro Plant Regeneration of Banana cv. Bhimkol (Musa balbisiana, BB) through Shoot Tip Culture

Bhimkol (Musa balbisiana, BB) is a wild diploid seeded cultivar, belonging to the family Musaceae. It is tolerance to pest and diseases, since it has high demand for commercialization for its high nutritive values. Therefore, the present study was undertaken to standardize the protocol for in vitro plant regeneration via shoot tips for further breeding programmes (in vitro mutagenesis). The efforts were made to improve the shoot proliferation efficacy through MS medium enhanced with different combinations of cytokinins (BAP and TDZ) and auxins (NAA) to achieve higher number of multiple shoots with shorter period. Among the different combinations of BAP, TDZ and NAA, T5 (MS + TDZ 0.1 mgL-1 + NAA 0.2 mgL-1) was recorded early response (6.60 days), percentage of explant response to initiation (93.33%), days taken for multiple shoot induction (46.40 days) and the highest number of shoots per explant (8.93). The days taken for rooting, number of roots per shoot and highest root length was noted in the treatment half strength MS + 1.0 mgL-1 IBA + 1.0 g L-1 activated charcoal.

Somatic embryogenesis of the selected intergeneric hybrid between Phalaenopsis 2166 and Vanda ‘Saint Valentine’: Application of NAA and TDZ

Abstract. Dwiati M, Susanto AH, Budisantoso I. 2022. Somatic embryogenesis of the selected intergeneric hybrid between Phalaenopsis 2166 and Vanda' Saint Valentine': Application of NAA and TDZ. Nusantara Bioscience 14: 160-165. Intergeneric hybridization between Phalaenopsis 2166 and Vanda' Saint Valentine' has produced a hybrid seedling with several characters potentially developing into plant individuals with flowers of better performance. Therefore, identical clones of the selected hybrid should be developed into PLBs using an in-vitro culture technique employing somatic embryogenesis supported by the application of plant growth regulators. This study aims to unveil the effect of NAA and TDZ in stimulating the formation of identical clones of the selected intergeneric hybrid between Phalaenopsis 2166 and Vanda' Saint Valentine'. The experiment was arranged in a factorial Randomized Complete Block Design (RCBD) involving two factors, i.e., types of plant growth regulators and the levels of concentrations of each substance. It was found that the combination of NAA and TDZ significantly affected the growth of the identical clones. Furthermore, the combination of NAA 0.5 mgL-1 and TDZ 1.5 mgL-1 resulted in clones that potentially differentiate into PLBs. This finding indicates that NAA and TDZ should be applied appropriately to stimulate somatic embryogenesis in the intergeneric hybrid.

Effect of Different Concentrations and Combinations of BAP and NAA on Micropropagation of Anthurium (Anthurium andraeanum Linn.) Red

Anthurium anderanum from Araneae family, is a plant with high commercial value. Tissue culture technique appears as an alternative way propagation to increase Anthurium production. The recent research intended to establish an efficient regeneration method for Anthurium andraeanum tissue culture. Young Anthurium plantlets including young shoots, petioles and leaves were applied for regeneration through in vitro indirect organogenesis method from the undefined plant tissues. Callus proliferation was induced by the use of Morishige and Skoog media along with 0.6 mg/l 2,4- Dichlorophenes acetic acid. Shoot regeneration medium consisted of MS salts with supplemented different combinations of BAP and NAA. The number of shoots per explants was significantly increased in 7 months of culture periods in all different combinations but there were found that 14 shoots/explant and 13.9 shoots/explant in NA+ BAP (0+3mg/l) and (0.2+0mg/l) respectively. The longest shoot length (17.3 cm) was found in NAA+BAP (0+0.5mg/l). Rooting was found automatically in shoot proliferation medium. Regenerated plantlets were potted in mixture of broken brick, coco husk and charcoal and successfully acclimatized.

Kombinasi BAP dan NAA untuk Media Perbanyakan Nanas Varietas Smooth Cayenne, Toboali in Vitro

Toboali pineapple variety is a smooth cayenne variety because it contains high antioxidants in the form of vitamins A and C. In addition, it is a source of citric and malic acids, substances that can be used to enhance the taste of the fruit. Pineapple is a plant that is propagated with vegetative organs such as suckers, slips, and crowns. However, conventional propagation has a low propagation rate. Tissue culture is an in vitro technique used in the rapid propagation of plants and in large quantities to meet the needs of cultivation and research. The selection of regeneration media is very influential on the development of plants in tissue culture. This study aimed to obtain the optimal media combination with BAP and NAA in the regeneration of Smooth Cayenne pineapple (Toboali) in vitro for mass propagation of seedlings and genetic engineering research. This study used 6 levels of BAP, namely 0, 1, 2, 3, 4, 5 mg.L<sup>-1</sup> and 4 levels of NAA, namely 0, 1, 2, 3 mg.L<sup>-1</sup>. The treatment used was a combination of BAP and NAA as many as 24 treatments with 3 replications. The best pineapple regeneration medium for mass plant propagation in this study was BAP 3 mg.L<sup>-1</sup> without NAA media which produced the highest shoots of 17.33 shoots per explant at 12 week after planting. The best pineapple regeneration medium for genetic engineering research was a combination BAP 3 mg.L<sup>-1</sup> and NAA 2 mg.L<sup>-1</sup> media which produced 18.33 shoots per explant from embryogenic callus.

In vitro Propagation of orchid (Dendrobium ovatum (L.) Kraenzl.) through Somatic Embryogenesis

An efficient in vitro regeneration protocol through somatic embryogenesis was established from longitudinally bisected protocorm (lTCL) of an endangered orchid Dendrobium ovatum. The efficiency of EC and SEs from the protocorms significantly relied on the concentration of PGRs. MS medium supplemented with TDZ (1.0 mg/l) induced optimum of EC (31.8%) and SEs (28.1/explant). Similarly, ZEA (0.5 mg/l) induced EC (27.6%) and SEs (17.1/explant). The combined treatment of TDZ (1.0 mg/l) and NAA (0.5 mg/l) resulted in the maximum induction of EC (58.6%) and SEs (39.8/explant) in an upright incubated explant. In another combined effect of ZEA (1.0 mg/l) and NAA (0.5 mg/l) induced EC (43.8%) and SEs (24.3/explant), whereas the combination of BAP (0.5 mg/l) and NAA (0.5 mg/l) produced EC (34.4%) and SEs (16.8/explants). The induced EC and SEs where healthier on medium containing TDZ + NAA than on the medium containing ZEA+NAA and BAP+NAA. The orientation of lTCL explants seemed to interact with position to affect the morphogenesis. The lTCL explants incubated in the upright orientation induced higher percentage of EC and a greater number of SEs/explants (EC-58.6% and SEs-39.8/explants) than that in the inverted orientation (EC- 16.4% and SEs-9.1/explants) irrespective of the PGRs. The SEs developed from the EC and the intermediate stage of PLBs were finally differentiated into plantlets. About 89% of plantlets were successfully acclimatized in the green house conditions. Plant Tissue Cult. & Biotech. 32(1): 53-66, 2022 (June)