Effects of polyamines, polyamine precursors, and polyamine biosynthetic inhibitors on somatic embryogenesis from eggplant (Solarium melongena) cotyledons

1988 ◽  
Vol 66 (9) ◽  
pp. 1734-1742 ◽  
Author(s):  
Pierre R. Fobert ◽  
David T. Webb

Eggplant (Solarium melongena L.) cotyledons grown on Murashige and Skoog medium with naphthaleneacetic acid formed callus, roots, and somatic embryos. Low levels of naphthaleneacetic acid (0.1 – 0.5 mg L−1) favoored rhizogenesis, intermediate levels (1.0 – 5.0 mg L−1) favoured embryogenesis, and high levels (10 – 50 mg L−1) favoured callogenesis. Addition of polyamines or their precursors did not induce morphogenesis on medium containing no growth regulator, nor did it affect embryogenesis on medium containing naphthaleneacetic acid, except at the highest concentrations tested, which were inhibitory. Enzyme-activated inhibitors of putrescine synthesis significantly reduced embryogenesis and stimulated rhizogenesis. α-Difluoromethylornithine was more potent in inhibiting embryogenesis and stimulating rhizogenesis than was α-difluoromethylarginine. α-Difluoromethylarginine did not inhibit growth and α-difluoromethylornithine stimulated growth. Addition of putrescine with α-difluoromethylornithine restored embryogenesis to control levels and reduced rhizogenesis. Competitive inhibitors of polyamine synthesis had nonspecific effects. Compared with seedling cotyledons, expiants grown on 5.0 mg naphthaleneacetic acid per litre contained slightly less free soluble putrescine and about the same amount of spermidine. At day 8, free putrescine and spermidine levels were higher in explants grown on naphthaleneacetic acid than in those grown on medium containing no growth regulator. Addition of α-difluoromethylornithine greatly reduced the putrescine and spermidine titres of the explants. Application of putrescine with α-difluoromethylornithine dramatically increased putrescine titres but not spermidine titres. Although the results suggest a role for polyamines in eggplant somatic embryogenesis, they do not support the hypothesis that polyamines act as auxin- or cytokinin-like growth regulators or as second messengers for auxin in this system.

2016 ◽  
Vol 71 (2) ◽  
Author(s):  
Fetrina OKTAVIA ◽  
. SWANTO ◽  
Asmini BUDIANI

SummaryTissue culture technique for arabica coffeefaces some problems, mainly in plantletsregeneration from cultured explants. Theobjectives of this experiment were to examine theeffect 2,4-D and 2-ip combinations on somaticembryogenesis and regeneration of arabicacoffee from several different explants. Basalmedium used in this experiment was MS mediumwith ½ concentration of macro and micro salts.Experiment to induce primary somatic embryos(SE) was arranged in factorial randomizedcomplete design with 10 repeats. The first factorwas the type of explants, leaf, epicotyl, hipocotyland root explants. The second factor was plantgrowth regulator i.e. combination of 1  M 2,4-Dwith 5, 10, 15, 20  M and combination of 5  M2,4-D with 5, 10, 15 and 20  M 2-ip. To multiplySE, secondary SE was induced from primary SEon medium containing combination of 0.6  MIAA and 13.3; 17.8 and 22.2  M BAP.Cotyledonary SE were germinated on mediacontaining GA 3 (0, 5, 10 and 15  M), and thenregenerated on medium free of growth regulator.Plantlets with 4-5 leaf pairs were transfered intothe soil medium for acclimatization. The resultsshow that primary SE can be induced from allexplants with the highest frequency on mediumcontaining 1  M 2,4-D and 15  M 2-ip.Induction of primary SE, in leaf explant wasmore effective than other explants. Mediumcontaining 0.6  M IAA and 22.2  M BAP gavethe highest percentage of SE multiplication i.e.52.6% with average SE number of 6.25. Plantletsregeneration can be conducted by culturing SEon maturation medium free of growth regulatorfor one month followed by germinating onmedium containing GA 3 , and then culturing onmedium free of growth regulator again. Thehighest percentage of germinated embryos wasobtained after three weeks and six weekscultured in the medium containing 5  M GA 3 , i.e49% and 90.15 respectively. From total plantletsobtained, 75% of them were normal. Sixtypercents of the young plants grew well in thegreenhouse.RingkasanTeknik kultur jaringan tanaman kopi arabikamasih menghadapi beberapa kendala terutamapada tingkat regenerasi planlet dari eksplan yangdikulturkan. Penelitian ini bertujuan untukmengetahui pengaruh kombinasi 2,4-D dan 2-ipterhadap embriogenesis somatik dan regenerasikopi arabika dari berbagai eksplan. Media dasaryang digunakan adalah medium MS ½konsentrasi garam makro dan mikro. Percobaaninduksi embrio somatik (ES) primer disusunmenurut rancangan acak lengkap faktorial dengan10 ulangan. Faktor pertama adalah jenis eksplan,erdiri atas daun, epikotil, hipokotil dan akar invitro. Faktor kedua adalah zat pengatur tumbuh,yaitu kombinasi 1 M 2,4-D dengan 5, 10, 15dan 20M 2-ip, serta kombinasi 5 M 2,4-Ddengan 5, 10, 15 dan 20 M 2-ip. Untuk mem-perbanyak jumlah ES yang didapatkan, dilakukaninduksi ES sekunder dari ES primer pada mediumyang mengandung kombinasi 0,6 M IAA dan13,3; 17,8 dan 22,2 M BAP. ES fase kotiledonkemudian dikecambahkan pada medium yangmengandung GA 3 (0, 5, 10 dan 15 M) danselanjutnya diregenerasikan pada medium tanpazat pengatur tumbuh. Planlet yang mempunyai4-5 pasang daun dipindahkan ke medium tanahuntuk aklimatisasi. Hasil yang diperolehmenunjukkan bahwa ES primer dapat diinduksipada semua eksplan yang digunakan denganfrekuensi tertinggi pada medium yang me-ngandung 1 M 2,4-D dan 15 M 2-ip. InduksiES primer pada eksplan daun lebih efektifdibandingkan eksplan lainnya. Untuk per-banyakan ES, medium yang mengandung IAA0,6 M dan BAP 22,2 M memberikanpersentase tertinggi pembentukan ES sekunderyaitu 52,6% dengan rata-rata jumlah ES 6,25.Regenerasi planlet dapat dilakukan denganmengkulturkan ES pada medium maturasi tanpazat pengatur tumbuh selama satu bulan, kemudiandikecambahkan dalam medium yang mengan-dung GA 3 , dan selanjutnya dipindah ke mediumtanpa zat pengatur tumbuh kembali.Perkecambahan ES tertinggi diperoleh padamedium dengan penambahan GA 3 5 M yaitu40,9% setelah tiga minggu dan 90,1% setelahenam minggu. Dari total planlet diperoleh 75%planlet normal. Hasil aklimatisasi menunjukkanbahwa 60% bibit mampu bertahan di rumah kaca.


HortScience ◽  
2014 ◽  
Vol 49 (12) ◽  
pp. 1558-1562 ◽  
Author(s):  
Yuyu Wang ◽  
Faju Chen ◽  
Yubing Wang ◽  
Xiaoling Li ◽  
Hongwei Liang

High-frequency somatic embryogenesis and plant regeneration were achieved from immature cotyledonary-stage embryos in the endangered plant, Tapiscia sinensis Oliv. Plant growth regulators with different concentrations and combinations on embryogenesis capacity were studied. The optimal explants for in vitro somatic embryogenesis were immature embryos in T. sinensis. A high callus induction rate of 100% was achieved on Murashige and Skoog (MS) basal medium supplemented with 1.0 mg·Ll−1 2,4-dichlorophenoxyacetic acid (2,4-D) and 0.5% (w/v) activated charcoal. Alternatively, a high induction rate (96.16%) of somatic embryogenesis was obtained on MS basal medium supplemented with the combination of 0.05 mg·L−1 α-naphthaleneacetic acid (NAA) and 0.2 mg·L−1 6-benzylaminopurine (6-BA), and somatic embryos proliferated fastest on the mentioned medium supplemented with 0.5% (w/v) activated charcoal and 3% (w/v) sucrose, inoculation of explants proliferating 21 times in the 23-day subculture. Of the 100 plantlets transferred to field after the acclimation, 95 (95%) survived. Based on the histocytological observations, the development of somatic embryos was similar to that of zygotic embryos. There were two accumulation peaks of starch grains in the embryogenic calli and in the globular-stage embryos, both closely related to the energy supply, and the embryoids were of multicelluar origin.


1990 ◽  
Vol 115 (4) ◽  
pp. 691-696 ◽  
Author(s):  
Rebecca M. Cade ◽  
Todd C. Wehner ◽  
Frank A. Blazich

Two studies were conducted to test the effects of various tissue culture media on somatic embryogenesis from cotyledon tissue of cucumber (Cucumis sativus L.). The two best media for embryo initiation were Murashige and Skoog (MS) salts and vitamins containing either 1 or 2 mg 2,4-D/liter and 0.5 mg kinetin/liter. In the second study, embryos developed more normally. More plantlets developed when tissue was removed from the initiation medium after 3 weeks and transferred to MS containing 1 mg NAA/liter and 0.5 mg kinetin/liter for 3 weeks, rather than leaving the embryos on a medium containing 2,4-D. Histological evidence indicated that the embryos were multicellular in origin. Charcoal in the maturation medium inhibited embryo development. Chemical names used: (2,4-dichlorophenoxy) -acetic acid (2,4-D); N-(2-furanylmethyl)-lH-purine-6-amine (kinetin); 1-naphthaleneacetic acid (NAA).


2018 ◽  
pp. 7-20
Author(s):  
Maria Daniela Artigas Ramirez ◽  
Rafael Fernandez Da Silva

Background and Aims: Meliaceae species are extremely recalcitrant during germination and in vitro processes. Therefore, this research focuses on characterization and optimization of a highly efficient system by secondary somatic embryogenesis in Azadirachta indica, which is an important step for enhancing secondary metabolite production and regeneration in recalcitrant species.Material and Methods: Leaf and cotyledon sections were induced in MS medium supplemented with benzylaminopurine (BAP) alone, or combined with 1-naphthaleneacetic acid (NAA) and, abscisic acid (BA) with thidiazuron (TDZ).Key results: Azadirachta indica developed primary somatic embryos with BAP. Shoot and root formation occurred at low concentrations of BAP, while somatic embryogenesis was favored under high levels of BAP or TDZ. Primary and secondary somatic embryos were evidenced continuously and asynchronously. The highest amount of somatic embryos was obtained with cytokinins. However, the concentration might be significant to differentiate between primary and secondary embryos. Moreover, the auxins are key for inducing histodifferentiation in embryos. Shoot induction occurred after transfer of the embryos to hormone-free MS medium. The shoots were rooted in MS1/2.Conclusions: The secondary somatic embryos were distinguished and characterized during the whole process and the efficient system was established with cotyledon sections at short term, which offers several advantages such as the production of metabolites.


Author(s):  
T.T.B. Phuong ◽  
V.P. Trung ◽  
N.H. An ◽  
N.D. Tuan ◽  
P.T.T. Nguyen

Background: Dinh Lang [Polyscias fruticosa (L.) Harms] is a medicinal plant widely grown in Vietnam, with proven note-worthy health benefits. However, Dinh Lang’s amounts of triterpenoid saponins could not meet the need of the pharmaceutical industry. Thus, this study’s purpose is to figure out the optimal condition for raising Dinh Lang’s cell biomass, rhizogenesis and somatic embryogenesis to provide materials for bioactive compound productions. Methods: Different 2,4-dichlorophenoxyacetic acid and α-naphthaleneacetic acid concentrations (0.5, 1.0, 1.5 and 2.0 mg/L) were examined to determine the best amount of each plant growth regulator for raising cells’ biomass, rhizogenesis and somatic embryogenesis. In each treatment, two grams of eight-week-old calli were cultured in 50 mL of liquid MS medium. Result: It is demonstrated by the results that liquid MS medium containing 1.5 mg/L α-naphthaleneacetic acid has the capacity of producing the highest numbers of somatic embryos (489 embryos per flask) and rooted cells (259.5 cells per flask), while the fresh weight of cells cultured in the medium given 1.5 mg/L 2,4-dichlorophenoxyacetic acid reached its peak of 5.7 g.


2016 ◽  
Vol 71 (2) ◽  
Author(s):  
Fetrina OKTAVIA ◽  
. SWANTO ◽  
Asmini BUDIANI

SummaryTissue culture technique for arabica coffeefaces some problems, mainly in plantletsregeneration from cultured explants. Theobjectives of this experiment were to examine theeffect 2,4-D and 2-ip combinations on somaticembryogenesis and regeneration of arabicacoffee from several different explants. Basalmedium used in this experiment was MS mediumwith ½ concentration of macro and micro salts.Experiment to induce primary somatic embryos(SE) was arranged in factorial randomizedcomplete design with 10 repeats. The first factorwas the type of explants, leaf, epicotyl, hipocotyland root explants. The second factor was plantgrowth regulator i.e. combination of 1  M 2,4-Dwith 5, 10, 15, 20  M and combination of 5  M2,4-D with 5, 10, 15 and 20  M 2-ip. To multiplySE, secondary SE was induced from primary SEon medium containing combination of 0.6  MIAA and 13.3; 17.8 and 22.2  M BAP.Cotyledonary SE were germinated on mediacontaining GA 3 (0, 5, 10 and 15  M), and thenregenerated on medium free of growth regulator.Plantlets with 4-5 leaf pairs were transfered intothe soil medium for acclimatization. The resultsshow that primary SE can be induced from allexplants with the highest frequency on mediumcontaining 1  M 2,4-D and 15  M 2-ip.Induction of primary SE, in leaf explant wasmore effective than other explants. Mediumcontaining 0.6  M IAA and 22.2  M BAP gavethe highest percentage of SE multiplication i.e.52.6% with average SE number of 6.25. Plantletsregeneration can be conducted by culturing SEon maturation medium free of growth regulatorfor one month followed by germinating onmedium containing GA 3 , and then culturing onmedium free of growth regulator again. Thehighest percentage of germinated embryos wasobtained after three weeks and six weekscultured in the medium containing 5  M GA 3 , i.e49% and 90.15 respectively. From total plantletsobtained, 75% of them were normal. Sixtypercents of the young plants grew well in thegreenhouse.RingkasanTeknik kultur jaringan tanaman kopi arabikamasih menghadapi beberapa kendala terutamapada tingkat regenerasi planlet dari eksplan yangdikulturkan. Penelitian ini bertujuan untukmengetahui pengaruh kombinasi 2,4-D dan 2-ipterhadap embriogenesis somatik dan regenerasikopi arabika dari berbagai eksplan. Media dasaryang digunakan adalah medium MS ½konsentrasi garam makro dan mikro. Percobaaninduksi embrio somatik (ES) primer disusunmenurut rancangan acak lengkap faktorial dengan10 ulangan. Faktor pertama adalah jenis eksplan,erdiri atas daun, epikotil, hipokotil dan akar invitro. Faktor kedua adalah zat pengatur tumbuh,yaitu kombinasi 1 M 2,4-D dengan 5, 10, 15dan 20M 2-ip, serta kombinasi 5 M 2,4-Ddengan 5, 10, 15 dan 20 M 2-ip. Untuk mem-perbanyak jumlah ES yang didapatkan, dilakukaninduksi ES sekunder dari ES primer pada mediumyang mengandung kombinasi 0,6 M IAA dan13,3; 17,8 dan 22,2 M BAP. ES fase kotiledonkemudian dikecambahkan pada medium yangmengandung GA 3 (0, 5, 10 dan 15 M) danselanjutnya diregenerasikan pada medium tanpazat pengatur tumbuh. Planlet yang mempunyai4-5 pasang daun dipindahkan ke medium tanahuntuk aklimatisasi. Hasil yang diperolehmenunjukkan bahwa ES primer dapat diinduksipada semua eksplan yang digunakan denganfrekuensi tertinggi pada medium yang me-ngandung 1 M 2,4-D dan 15 M 2-ip. InduksiES primer pada eksplan daun lebih efektifdibandingkan eksplan lainnya. Untuk per-banyakan ES, medium yang mengandung IAA0,6 M dan BAP 22,2 M memberikanpersentase tertinggi pembentukan ES sekunderyaitu 52,6% dengan rata-rata jumlah ES 6,25.Regenerasi planlet dapat dilakukan denganmengkulturkan ES pada medium maturasi tanpazat pengatur tumbuh selama satu bulan, kemudiandikecambahkan dalam medium yang mengan-dung GA 3 , dan selanjutnya dipindah ke mediumtanpa zat pengatur tumbuh kembali.Perkecambahan ES tertinggi diperoleh padamedium dengan penambahan GA 3 5 M yaitu40,9% setelah tiga minggu dan 90,1% setelahenam minggu. Dari total planlet diperoleh 75%planlet normal. Hasil aklimatisasi menunjukkanbahwa 60% bibit mampu bertahan di rumah kaca.


HortScience ◽  
1992 ◽  
Vol 27 (1) ◽  
pp. 63-65 ◽  
Author(s):  
Les Frey ◽  
Yehoshua Saranga ◽  
Jules Janick

Somatic embryogenesis was induced from internodal callus of `Scania', `Improved White Sim', and `Sandra' carnation (Dianthus caryophyllus L.). The optimum protocol for the induction of somatic embryogenesis included initiation of callus in liquid basal Murashige and Skoog medium supplemented with 3.0 μm 2,4-D followed by transfer to liquid basal medium lacking 2,4-D for embryo development. Somatic embryos originated from single cells and early embryonic development proceeded conventionally (i.e., via globular, heart-shaped, and torpedo stages), but clearly developed apical or root meristems were not always formed. A few embryos developed into seedlings and were acclimatized to ex vitro conditions. Chemical name used: 2,4-dichlorophenoxyacetic acid (2,4-D).


Author(s):  
Fitria Ardiyani ◽  
Edy Setiti Wida Utami ◽  
Hery Purnobasuki

Coffea liberica is a variety of coffee that tolerant to marginal land, especially peatlands. One of propagation methods in C. liberica is somatic embryogenesis(SE) which producing large number of true-to-type plant seedlings in a short time. This research aimed at studying the effect of application of plant growthregulator (PGR) on quality and weight of somatic embryo of C. liberica. Somatic embryo in development stage was induced by Murashige and Skoog medium containing cytokinin as benzyl amino purin (BAP) and auxin as 2,4-dichlorophe-noxyacetic acid (2,4-D). While cotyledonary embryo in germination stage was induced by Murashige and Skoog medium containing cytokinin (BAP) and auxins as 2,4-D, indole acetic acid (IAA) and naphthaleneacetic acid (NAA). The resultsshowed that the application of auxins and cytokinins on development stage affected the formation of embryos, texture of calli, color of calli and embryos, and weight of somatic embryo. It also influenced the shoot and root formation, color and weight of geminating embryos of C. liberica at the germinating stage. During the development stage, addition of 1 mg/L BAP in the absence of 2,4-D in MS medium produced the highest quality of somatic embryo of C. liberica. This medium also produced heaviest somatic embryos but with lighter callus. While in germination stage, all medium treatments produced a typical germinating embryo. Coffea liberica germinating embryo growth optimally on MS medium containing 0.5 mg/L BAP as a single chemical or 0.5 mg/L BAP in combination with 0.5 mg/L IAA for shooting development. Whereas on rooting development, addition of 0.5 mg/L NAA on MS medium produced an optimal germinating embryo. Moreover, germination embryo of C. liberica recorded the highest in terms of dry weight on MS media with addition of 0.5 mg/L BAP. Application of appropriate concentration of auxin and cytokinin is needed to support the formation of somatic embryo and germinating embryo.


2014 ◽  
Vol 66 (2) ◽  
pp. 159-163 ◽  
Author(s):  
Alina Trejgell ◽  
Andrzej Tretyn

Immature embryos of <em>Pharbitis nil</em> Chois. were used for the study. They were isolated from previously sterilized fruits and afterwards transferred to Murashige and Skoog (MS) growth medium with 0.8% agar. Immature embryos were cut across their axis. After 6-8 weeks of cultivation in the injury place (hypocotyl-root region) somatic embryos were formed. These embryos were isolated and each of them was transferred into a separate tube containing MS supplemented with naphthaleneacetic acid (NAA; in concentration 0.1 mg•dm<sup>-3</sup> ) and gibberellic acid (GA<sub>3</sub>; in concentration 0.5 mg•dm<sup>-3</sup>). Under these conditions about 26% of somatic embryos regenerated into complete plants. Two-three weeks after the photoperiodic induction flowers appeared on these plants. A few weeks after pollination normal seeds were developed from these flowers.


HortScience ◽  
2000 ◽  
Vol 35 (4) ◽  
pp. 568B-568a
Author(s):  
Lianghong Chen ◽  
Ajmer S. Bhagsari ◽  
Soon O. Park ◽  
Sarwan Dhir

This study was carried out to optimize conditions for plant regeneration of sweetpotato [Ipomoea batatas (L.) Lam] using shoot tips, petioles, and leaves of Selection 75-96-1 as explants in Murashige and Skoog (MS) with several growth regulators at different levels. Callus initiation and callus proliferation media were 9.0 μm 2,4-dichlorophenoxyacetic acid (2,4-D) and 9.0 μm 2,4-D + 1.1 μm N6-benzyladenine (6-BA) in protocol I; 8.1 μm α-naphthaleneacetic acid (NAA) + 1.2 μm kinetin (KIN) and 5.4 μm NAA + 4.6 μm KIN in protocol II; 0.9 μm 2,4-D, and 0.9 μm 2,4-D + 1.2 μm N-isopenylamino purine (2iP) in protocol III; NAA (8.1 μm) + KIN (1.2 μm) and 2,4-D (0.9 μm) + 2ip (1.2 μm) in protocol IV, respectively. In protocol I and II, shoot tip, petiole, and leaf were used, but only petiole and leaf in protocol III and IV. In the protocol I and II, somatic embryos were obtained only from shoot tip explants; in protocol III and IV, only from petioles. The frequencies of somatic embryo development were 33.3% in protocol I, 42.1% in protocol II, 21.2% in protocol III, and 10.3% in protocol IV, respectively. The leaf explants failed to produce somatic embryos in all the experiments. In protocol I, somatic embryogenesis occurred through the well-known sequence of globular-, heart-shaped-, torpedo-, and cotyledon-type embryos. However, in protocol II, the structures resembling plumule and radicle were observed before the emergence of torpedo/cotyledon type embryo clusters. The somatic embryogenesis in protocol III and IV was similar to that in protocol I. Growth regulators influenced somatic embryo development. Further, this study showed that explant resource and growth regulators affected the frequency of plant regeneration in sweetpotato.


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