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Micro Propagation and Organogenesis of Peperomia obtusifolia
Abstract
Peperomia obtusifolia is a bushy upright species belong to the Piperaceae, native to Florida, Mexico and the Caribbean, as an alternative to the conventional vegetative propagation methods is becoming an important way to increase shoot proliferation rates and produce of new varieties through transformation; therefore, the purpose of this study was to establish a protocol for in vitro production of Peperomia obtusifolia and to compare between two explants for shoot proliferation and organogenesis. Single node stem segments and leaf segments of Peperomia obtusifolia were cultured on solidified Murashige and Skoog (MS) basal medium supplemented with kinetin (Kin) at 0, 2.5, 5 and 10 mg L-1 and naphthalene acetic acid (NAA) at 0, 1, 2.5 and 5 mg L-1. All explants generated from leaf segments turned dark brown and died no shoots or callus were formed. Percentage of successful shoot regenerated from single node stem segments increased by increasing kinetin concentration then declined at the highest concentration of kinetin (10 mg L-1) while different concentrations of NAA almost had no effect on percentage of shoot formation. Kinetin at 5 mg L-1 combined with NAA at 1 mg L-1 was the most effective concentrations in percentage of shoot proliferation, number of shoots, fresh weight, total leaf area, total chlorophyll and callus weight on the other hand this concentration caused a reduction in shoot length and a delay in days to proliferation.
... combination with high cytokinins concentration promoted shoot proliferation in Crambetataria(Piovan et al. 2011)andP. Obtusifolia(Hany and Amira 2014)belongs to the family Piperaceae. In vitro propagation is an excellent technique to obtain maximum number of plants within a short period of time . ...
... There are few studies conducted on Genus Peperomia for propagation but none of them specifically tested for P. pellucida. Naggar and Osman (2014) reported micropropagation of Peperomia obtusifolia using BA and KN media where KN (5 mg L -1 ) combined with α-Naphthalene Acetic Acid (NAA) (1 mg L -1 ) displayed the most effective concentrations for shoot proliferation, number of shoots, fresh weight, total leaf area, total chlorophyll and callus weight. For P. obtusifolia KN was inefficient compared to Benzylaminopurine (BAP) but MS media supplemented with the later resulted in multiple adventitious root development (Al-Ahmad 2015). ...
... Means followed by the same letters in each column are not significantly different (P = 0.05) using Duncan's multiple range test 1.0 mg/l in combination with 0.7 mg/l NAA responded highest shoot regeneration (89 %) with highest number of shoots (25.57) and shoot length (3.87) on leaf derived callus (Table 3). Similar results have also been observed in leaf explants of in Peperomia obtusifolia (Naggar and Osman 2014) and Ziziphora tenuior (Dakah et al. 2014). Likewise, internodal explants cultured on MS medium supplemented with KN (1.0 mg/l) and NAA (0.7 mg/l) showed higher regeneration frequency (85 %) and highest number (16.42) of shoots and shoot length (2.94). ...
The present study describes the plant propagation via indirect organogenesis from in vitro derived leaf and internode explants of Plectranthus bourneae, an endemic plant to south India. Leaf and internodal explants successfully callused on Murashige and Skoog medium (MS) supplemented with different concentrations of auxins [2,4-D (2,4-dichlorophenoxyacetic acid), NAA (α-naphthalene acetic acid), IAA (indole-3 acetic acid), IBA (indole-3-butyric acid) and PIC (Picloram); 0.1–2.0 mg/l] in combination with BA (6-benzyladenine) (0.5 mg/l). Maximum callus induction (98 %) was achieved from leaf explant followed by internodal explant (89 %) at 1.0 mg/l NAA, 0.5 mg/l BA. Leaf derived callus showed better shoot regeneration (29.71 shoots) on MS medium containing 1.0 mg/l KN (kinetin), 0.7 mg/l NAA, and 50 mg/l CH (casein hydrolysate) followed by internodal callus (19.71). A maximum of 19.14 roots/shoot was observed at 1.0 mg/l IBA. The rooted plantlets were successfully hardened and transferred to greenhouse condition with 80 % survival. This system could be utilized for large-scale multiplication of P. bourneae by tissue culture.
Kegiatan penelitian dilaksanakan pada bulan Maret–Juli 2021 di Bogor, Jawa Barat. Penelitian bertujuan untuk meningkatkan kemampuan dalam pengelolaan tanaman hias khususnya peperomia, bromelia, begonia, dan hortensia. Kegiatan pengelolaan tanaman hias mencakup jadwal produksi mulai dari persiapan tanam hingga siap panen dan kriteria panen tanaman. Pengamatan tanaman Peperomia obtusifolia, Neoregelia schultesiana, dan Hydrangea macrophylla meliputi jumlah daun dan tinggi tanaman. Pengukuran tanaman Peperomia argyreia dan Peperomia caperata meliputi jumlah tunas. Pengukuran Begonia ‘orococo’ meliputi panjang daun, lebar daun, dan jumlah daun. Kegiatan budidaya tanaman hias yang dilakukan meliputi persiapan media tanam, pembibitan, penyiraman, pemupukan, pengendalian hama penyakit, panen, dan pascapanen. Hasil pengamatan pada tanaman Peperomia obtusifolia, Neoregelia schultesiana, dan Begonia ‘orococo’ yang menggunakan metode perbanyakan stek batang tergolong cepat, sedangkan Peperomia caperata dan Peperomia argyreia yang diperbanyak dengan menggunakan metode stek daun memiliki pertumbuhan yang lambat sehingga selama penelitian tanaman tidak memenuhi kriteria pemasaran. Hydrangea macrophylla memiliki umur panen rata-rata sekitar 15 MST. Kemampuan teknis mahasiswa meningkat dalam pengelolaan tanaman hias. Hasil analisis usahatani dengan asumsi luasan 200 m2 pada Peperomia obtusifolia, Peperomia argyreia, Peperomia caperata, Begonia ‘orococo’, dan Neoregelia schultesiana menunjukkan hasil R/C rasio >1 yang berarti menguntungkan. Kata kunci: analisis usaha tani, kriteria panen, pembibitan tanaman hias, stek daun, stek batang
Medicinal herbs are the main source of bioactive compounds used in the medical industry. White squill (Urginea maritima) is an important medicinal and ornamental plant cultivated in the Mediterranean region. This study reports an efficient protocol for in vitro propagation of Urginea and investigates important bioactive compounds present in the bulbs and in vitro–produced callus. The least number of days for callus induction and shoot regeneration was achieved with Murashige and Skoog (MS) basal media supplemented with 1.0 mg L−1 1-naphthaleneacetic acid (NAA) plus 0.1 mg L−1 6-benzylaminopurine (BAP) and 1.0 mg L−1 NAA plus 0.4 mg L−1 BAP, respectively, while the highest number of shoots and fresh weight were obtained at medium supplemented with 1.0 mg L−1 NAA plus 0.5 mg L−1 BAP. Gas chromatography-mass spectrometry (GC–MS) analysis of Urginea bulb methanol extract showed the existence of important secondary metabolites, such as palmitic acid (C16H32O2), 9-hexadecenoic acid (C16H30O2), phthalic acid 2-ethylhexyl propyl ester (C19H28O4), tetradecanoic acid (C14H28O2), undecanoic acid (C11H22O2), and oleic acid (C18H34O2), in addition to other important compounds, such as 13-heptadecyn-1-ol, 9,12-octadecadienoic acid, 1-monolinoleoylglycerol trimethylsilyl ether, 2-methyl-1-hexadecanol, and octadecanoic acid. Callus methanol extracts showed a reduction in the percentages of most phyto-components compared to bulb extract except for oleic acid, 3-(octadecyloxy) propyl ester and 3-hydroxydodecanoic acid; on the other hand, some important compounds were detected only in callus extract possessing anti-cancer, antiviral, and anti-inflammatory effects, such as farnesol (C15H26O), 7-methyl-Z-tetradecen-1-ol acetate (C17H32O2), ethyl iso-allocholate (C26H44O5), 4-trifluoroacetoxypentadecane (C17H31F3O2), and 2-hydroxyhexadecanoic acid (C16H32O3).
Peperomia albovittata and P. galioides are two herbaceous species, belonging to the Piperaceae family, collected in the Andean district of Kañaris (1128 to 1550 m above sea level), in Lambayeque region, Perú. These species are recalcitrant to propagation by conventional vegetative methods, whereby the present study aims to in vitro propagate of Peperomia albovittata and P. galioides through by organogenesis. Single nodes with axillar buds, stem segments (internodes), leaves and petioles were cultured on gellified basal culture medium and was added IAA-GA3, NAA-GA3 and NAA-BAP at different concentrations. Only in the NAA-GA3 combination clonal propagation from nodal segments and direct organogenesis in internodes were observed. Callus induction and proliferation was observed in some interactions of NAA-BAP, although all explants from leaves turned dark brown and died. On the other hand, shoot regeneration was observed only in P. galioides. Both species, especially P. albovittata, showed a high degree of recalcitrance to the induction of in vitro morphogenic processes.
The effect of benzylaminopurine (BAP) and kinetin alone and in combination with indole 3-acetic acid (IAA) on shoot proliferation of 'Basrai' (Musa spp., AAA group) was investigated. Shoot tips (4-6 mm) were excised from field-grown suckers to initiate the cultures. Concentrations of BAP and kinetin ranged from 0.0 to 8.0 mg·L -1 each on solid or in liquid MS medium. When liquid medium was used, cultures were agitated continuously on an orbital shaker moving at 40 rpm. Three subculture regimes were employed; after each subculture, the number of shoots regenerated from each explant was counted. The results showed that the multiplication rate was significantly (P ≤ 0.05) dependent upon cytokinin type, its concentration, and type of medium used. The maximum number of shoots regenerated from a single shoot tip was achieved in liquid MS medium containing 4.0 mg·L -1 BAP. There was no significant difference between liquid and solid medium when kinetin was used; however, kinetin at 4.0 mg·L -1 or above yielded significant results as compared with the control and lower kinetin concentrations. The results demonstrated that 4.0 mg·L -1 BAP with 1.0 mg·L -1 IAA in liquid medium was best for shoot multiplication and shoot height during micropropagation of 'Basrai'.
Several species in the genus Peperomia Ruiz & Pav. have giant chloroplasts in the palisade parenchyma of their leaves. Because of this unique feature, Peperomia metallica L. Linden & Rodigas has become a valuable model plant for studying plastid biology. However, the use of Peperomia for molecular studies has been limited by the lack of efficient regeneration and transformation protocols. In this work, we aimed to develop a reliable regeneration system for Peperomia from leaf explants. Using MS medium complemented with different combinations of phytohormones, a highly efficient in vitro regeneration protocol was developed for P metallica and P peduncularis, another species with giant chloroplasts. Regeneration occurs through direct organogenesis from leaf discs at a very high frequency. Our protocol can provide a reliable tissue culture system for future Peperomia genetic transformation experiments.
The relative stabilities of IAA and IBA under various tissue culture procedures were determined. IBA was significantly more stable than IAA to autoclaving. IBA was also found to be more stable than IAA in liquid Murashige and Skoog medium (MS) under growth chamber conditions. The stabilities of IBA and IAA were similar in agar-solidified MS. Light provided by cool-white fluorescent bulbs promoted degradation of IAA and IBA in both liquid and agar media. Activated charcoal in concentrations as high as 5% was found to adsorb more than 97% of IAA and IBA in liquid MS. These results have important implications for the preparation, storage, and handling of IBA and IAA in plant tissue culture. Chemical names used: indole-3-acetic acid (IAA); indole-3-butyric acid (IBA).
Objective
This is the first attempt for an efficient plant regeneration protocol through in vitro direct organogenesis for a valuable medicinal plant, Dipteracanthus prostratus using nodal segment.Methods
Multiple shoots were induced from nodal explants cultured on Murashige and Skoog medium supplemented with kinetin (KIN), 6-benzylaminopurine (BAP) and 1-Naphthaleneacetic acid (NAA).ResultsMaximum shoot responses (80%) were obtained with kinetin at 1.0 mg−1. The rate of shoot multiplication was maintained in subsequent subculture on similar fresh culture medium. The highest shoot length (3.96cm) was obtained with seventy three percentages of shoots at 0.2 mg−1 NAA along with 1.0 mg−1 kinetin. Maximum length of root (3.63cm) was formed at 0.5 mg−1 IBA with significant responses (80%). Rooted plantlets were then transferred to perforated plastic cups and grown in the green house at 80% survival rate.Conclusions
The highest survival rate was noticed and this plant developmental protocol could be used for large— scale regeneration of D. prostratus.
Despite plant age is a key factor to understand the performance and survival of any given species in the field, little is known about its influence on the endogenous levels of cytokinins in juvenile individuals of perennial plants. In this study, we hypothesized that plant age in juvenile mastic trees may affect cytokinin levels and lead to significant changes in cytokinin-related processes, such as leaf growth and pigment accumulation. Two groups of juvenile plants aged 2 and 5 years old were grown under Mediterranean field conditions to evaluate the age-related differences in the endogenous levels of cytokinins and auxins, leaf biomass and area, pigments levels and the efficiency of PSII photochemistry (Fv/Fm ratio). Furthermore, regression analyses between cytokinin levels and several indicators of stress performance were carried out in a population of juvenile mastic trees. Results showed that endogenous levels of cytokinins, particularly 2-isopentenyladenine (2-iP), increased during periods of intense leaf growth. However, plant age did not affect 2-iP levels, but those of trans-zeatin (Z); 2-year-old plants showing 2-fold higher levels of Z compared to 5-year-old plants, the latter having 11-fold higher total plant biomass. Plant age-related reductions in cytokinins were not associated with reductions in leaf growth in the largest individuals, but with an increased chlorophyll (Chl) a/b ratio. Further regression analyses revealed that although enhanced Chl a/b ratios predispose leaves to suffer photooxidative stress, as indicated by increased anthocyanins levels and lowered Fv/Fm ratios in juvenile mastic trees, individuals of different ages showed a similar degree of stress tolerance. It is concluded that plant age may alter the endogenous composition of cytokinins and the pattern of pigment accumulation in leaves, but this does not reduce leaf growth or stress tolerance in this species.
In vitro bulb scales of Lilium longiflorum ‘Georgia’ were cultured on MS medium with BA (benzyladenine) to induce shoots. After 6 weeks, the frequency of shoot formation was very high (more than 92.3%) on the media with 0.4–8.9 μM BA. Medium supplemented with 2.2 μM BA was most effective in inducing shoots from bulb scales. The formed shoot clusters were cut into sections of about 5–7 mm in size at random, and then cultured on shoot proliferation medium. The proliferation of shoots was also achieved very efficiently on medium with 2.2 μM BA and 2.9 μM IAA (indole acetic acid) (more than 9 shoots/culture). The addition of liquid medium into the same vessels was performed to form normal bulblets, after shoots were proliferated on the media with 2.2 μM BA and 2.9 μM IAA for 8 weeks. The addition of liquid medium stimulated the formation and growth of bulblets remarkably, compared with no addition of liquid medium. The liquid medium supplemented with 5.0–10.0 g/L activated charcoal and 250 g/L sucrose was most effective on the formation and growth of in vitro bulblets from shoot clusters. Thirty milliliters of liquid medium added into the vessels was suitable for the formation and growth of bulblets from shoot clusters. The formation and growth of bulblets were not affected by light. The formed bulblets grew normally, and some bulblets generated flower stems in greenhouse after cold treatment at 5 s for 2 months.