Analysis of ancient DNA from in vitro grown tissues of 1600-year-old seeds revealed the species as Anagyris foetida

2012 ◽  
Vol 22 (4) ◽  
pp. 279-286 ◽  
Author(s):  
Murat Özgen ◽  
Aslı Özdilek ◽  
Melahat A. Birsin ◽  
Sertaç Önde ◽  
Derya Şahin ◽  
...  
Keyword(s):  
Nuclear Dna ◽  
Adventitious Shoots ◽  
Its Region ◽  
Naphthaleneacetic Acid ◽  
Archaeological Excavation ◽  
Relict Species ◽  
Western Turkey ◽  
Callus Tissues ◽  
Seed Characteristics

AbstractSeven ancient seeds, about 1600 years old, were found during an archaeological excavation in Asar Island which is located in south-western Turkey. These seeds were subjected to germination, in vitro callus induction and molecular characterization experiments to test the viability and plant origin of the seeds. Six of the seven seeds had viable seed components (such as cotyledons) and produced callus tissue in Murashige and Skoog medium supplemented with 2 mg l− 1 6-benzylamino purine (BAP), 0.2 mg l− 1 1-naphthaleneacetic acid (NAA), 20 g l− 1 sucrose, 2 mg l− 1 glycine and 7 g l− 1 agar, but the calli from these seeds failed to yield adventitious shoots. DNA samples from callus tissues produced by ancient seeds in vitro were of good quality. The internal transcribed spacer (ITS) region in nuclear DNA (nDNA) of ancient seeds was amplified successfully. The sequences from amplified ITS DNA products of six ancient seeds indicated that their ITS sequences matched those of Anagyris foetida after subjecting them to BLAST searches in international sequence databases (NCBI). A. foetida is a relict species endemic to the Mediterranean region and used as a herbal medicine. We believe that seed characteristics such as the very hard, extremely smooth and shiny testa, toxic anagyrine alkaloid content and their storage in a pot further improved the longevity of these ancient seeds.

Introduction of Eucommia ulmoides Oliv. to in vitro Culture

2020 ◽  
pp. 38-50
Author(s):  
A.V. Zhigunov ◽  
◽  
Q.T. Nguyen
Keyword(s):  
Herbal Medicines ◽  
Naphthaleneacetic Acid ◽  
Eucommia Ulmoides ◽  
Juvenile Period ◽  
Dioecious Species ◽  
Relict Species ◽  
Mass Reproduction ◽  
Special Value ◽  
The Republic

The increasing need for herbal medicines requires the study of not only biological resources of medical plants, but also methods for their reproduction. Of special value are the medicinal plants that have a long history of success in traditional medicine. One of such plants is Eucommia ulmoides Oliv., which belongs to a rare relict species growing in natural conditions, for the most part, in the undergrowth of humid subtropical forests in China, mainly in the middle course of the Yangtze river. E. ulmoides compares favorably with most subtropical plants owing to its significant frost resistance, which makes it possible to cultivate it outside the humid subtropics. It has been widely introduced in Krasnodar Krai and in the Republic of Adygea (Russia) since the mid-20th century and successfully adapted to various environmental conditions in the Northwest Caucasus. The increasing demand for E. ulmoides bark can only be satisfied by laying out industrial plantations. However, the difficulties encountered in the traditional seed reproduction of E. ulmoides (dioecious species, pollen low quality, parthenocarpy, prolonged seed dormancy, irregular fruiting, long juvenile period, etc.) make scientists turn to modern biotechnological methods of plant propagation. While considering cultivation of planting material, we should focus on highly efficient methods that ensure stable and mass reproduction of the plants under study. An important role is played here by in vitro plant regeneration. The effectiveness of biotechnology methods is due to a reduction in timing of obtaining a large number of vegetative progeny of plants difficult for propagation, as well saving of the area required for their cultivation. The conditions for producing an aseptic culture of E. ulmoides were chosen based on the results of the studies. The highest degree of sterilization of E. ulmoides shoot segments was achieved when the explants were sequentially immersed first in 70 % ethanol (30 s) and then in 0.1 % mercuric chloride solution (5 min). With such a sterilization procedure, 63.3 % of the studied cuttings were made sterile, and 56.7 % of them proved to be viable. The optimal composition of the nutrient medium for regeneration of E. ulmoides microshoots has been determined: MS medium complemented with 1 mg/L 6-Benzylaminopurine (BAP) + 0.2 mg/L 1-Naphthaleneacetic acid (NAA). The best media for explant rooting are the following: 2/3 MS + 1.5 mg/L NAA + 30 g sucrose + 7 g agar; 2/3 MS + 1 mg/L NAA + 0.4 mg/L IBA + 30 g sucrose + 7 g agar.

In vitro propagation of Crambe maritima

1987 ◽  
Vol 65 (1) ◽  
pp. 72-75 ◽  
Author(s):  
J. Y. Peron ◽  
E. Regnier
Keyword(s):  
In Vitro Propagation ◽  
Indoleacetic Acid ◽  
Adventitious Shoots ◽  
Basal Medium ◽  
Naphthaleneacetic Acid ◽  
Petiole Explants ◽  
Acclimatization Period

A method for rapid micropropagation of sea kale (Crambe maritima L.) was developed. Petiole explants placed in vitro on a medium containing 0.5 mg/L indoleacetic acid (IAA), 6.0 mg/L kinetin, and 1.5 mg/L benzylaminopurine developed callus within 15 days and shoots within 28 days. Nearly four adventitious shoots could be developed within 3 weeks by placing the initial shoot on media without IAA. To develop roots, the shoots were then transferred to the basal medium containing 0.1 to 1.0 mg/L indolbutyric or α-naphthaleneacetic acid. Rooted plantlets were obtained within 2 or 3 weeks. After an acclimatization period of 6 weeks in a greenhouse in unsterilized medium, the plantlets could be set outdoors.

scholarly journals A Highly Efficient In Vitro Cranberry Regeneration System Using Leaf Explants

HortScience ◽  
2000 ◽  
Vol 35 (5) ◽  
pp. 948-952 ◽  
Author(s):  
Luping Qu ◽  
James Polashock ◽  
Nicholi Vorsa
Keyword(s):  
Growth Regulators ◽  
Adventitious Shoots ◽  
Basal Medium ◽  
Naphthaleneacetic Acid ◽  
Leaf Position ◽  
Regeneration System ◽  
Abaxial Side ◽  
Adventitious Regeneration ◽  
Explant Orientation

A very efficient adventitious regeneration (shoot organogenesis) system for cranberry (Vaccinium macrocarpon Ait.) leaves was developed. A basal medium consisting of Anderson's rhododendron salts and Murashige and Skoog's (MS) organics, supplemented with 10.0 μm thidiazuron (TDZ) and 5.0 μm 2ip, was effective for adventitious regeneration from leaves for the five cranberry cultivars tested: `Early Black', `Pilgrim', `Stevens', `Ben Lear', and `No. 35'. Parameters examined included: 1) varying combinations of three plant growth regulators (TDZ, 2ip, and NAA); 2) explant orientation (adaxial vs. abaxial side in contact with the medium); and 3) leaf position relative to the apical meristem from the source plant. Cultivars varied in regeneration frequency, but cultivar × growth regulator interaction was nonsignificant. With optimal treatment conditions, regeneration occurred on more than 95% of the explants, with `Early Black' and `Pilgrim' producing as many as 100 shoot meristems per explant. At all concentrations tested, NAA (as low as 0.1 μm) increased callus formation and significantly reduced regeneration. Emerging adventitious shoots were always observed on the adaxial side of the leaves regardless of explant orientation on the medium. Regeneration was much greater when the abaxial side was in contact with the medium, and was not related to leaf position on the source plants. Elongation of adventitious shoots began ≈2 weeks after transfer to the basal medium without growth regulators. Cuttings of elongated shoots rooted 100% both in vitro in the basal medium and ex vitro in shredded sphagnum moss. The high regeneration efficiency achieved by using this system will be very useful in the application of techniques, such as Agrobacterium- and particle bombardment-mediated transformation. Chemical names used: 1-phenyl-3-(1,2,3-thiadiazol-5-yl) urea (thidiazuron, TDZ); N6-(γ-γ-dimethyallylamino) purine (2ip); α-naphthaleneacetic acid (NAA).

scholarly journals Adventitious Shoot Regeneration and Rooting of Prunus serotina In Vitro Cultures

HortScience ◽  
2006 ◽  
Vol 41 (1) ◽  
pp. 193-201 ◽  
Author(s):  
Ana Carolina Espinosa ◽  
Paula M. Pijut ◽  
Charles H. Michler
Keyword(s):  
Shoot Regeneration ◽  
Adventitious Shoots ◽  
Leaf Explants ◽  
Adventitious Shoot ◽  
Prunus Serotina ◽  
Naphthaleneacetic Acid ◽  
Eastern United States ◽  
Continuous Darkness ◽  
Number Of Shoots

A complete regeneration protocol was developed for Prunus serotina Ehrh., an important hardwood species for timber and sawlog production in the central and eastern United States. Nodal sections were cultured on Murashige and Skoog (MS) medium supplemented with 4.44 μm 6-benzylaminopurine (BA), 0.49 μm indole-3-butyric acid (IBA), and 0.29 μm gibberellic acid (GA3). In vitro leaf explants of three genotypes were placed on woody plant medium (WPM) supplemented with 0, 2.27, 4.54, or 6.81 μm thidiazuron (TDZ) in combination with 0, 0.54, 1.07, or 5.37 μm naphthaleneacetic acid (NAA), and on WPM supplemented with 0, 4.44, 8.88, or 13.32 μm BA in combination with 0, 0.54, 1.07, or 5.37 μm NAA. Cultures were maintained either in continuous darkness for 5 weeks, or in the dark for 3 weeks and then transferred to a 16-hour photoperiod. TDZ and the genotype had a significant effect on the number of shoots regenerated. The maximum mean number of shoots regenerated per explant (5.05 ± 1.14) was obtained with 2.27 μm TDZ plus 0.54 μm NAA with the 3-week dark period then light treatment. The highest percent shoot regeneration (38.3) and mean number of shoots (4.13 ± 0.97) was obtained with 6.81 μm TDZ plus 1.07 μm NAA. The highest rooting (27%) of adventitious shoots and number of roots per shoot (2.3 ± 0.2) was obtained with 2.5 μm IBA when shoots were maintained for 7 days in the dark on rooting medium before transfer to a 16-hour photoperiod. The highest rooting (70%) of nodal explant-derived stock cultures and number of roots per shoot (2.7 ± 0.9) was also obtained with 2.5 μm IBA, but when shoots were maintained for 4 days in the dark before transfer to a 16-hour photoperiod. In total, 86% of the plantlets survived acclimatization to the greenhouse and 100% survival after overwintering in cold-storage.

scholarly journals In vitro organogenesis in some citrus species

2011 ◽  
Vol 33 (2) ◽  
pp. 526-531 ◽  
Author(s):  
Evandro Henrique Schinor ◽  
Fernando Alves de Azevedo ◽  
Francisco de Assis Alves Mourão Filho ◽  
Beatriz Madalena Januzzi Mendes
Keyword(s):  
Culture Medium ◽  
Culture Media ◽  
Adventitious Shoots ◽  
Medium Composition ◽  
Naphthaleneacetic Acid ◽  
In Vitro Organogenesis ◽  
Basal Culture Medium ◽  
Medium Supplementation ◽  
Media Supplementation

In vitro organogenesis of Citrus was studied for the genotypes Citrus sinensis cv. 'Natal', C. limonia, C. volkameriana, and C. aurantium, with the use of epicotyl segments-derived explants, cultured in MT salts and vitamins medium supplemented with different concentrations of 6-benzylaminopurine (BAP - 0.0; 0.5; 1.0; 1.5 or 2.0 mg L-1). For the recalcitrant genotypes C. limonia and C. aurantium the in vitro organogenesis was also studied with internodal segments-derived explants, cultured in MT salts and vitamins medium supplemented with 0; 0.5; 1.0; 2.0, or 4.0 mg L-1 of BAP. The efficiency of culture medium supplementation with the combination of BAP (0.0; 1.0, or 2.0 mg L-1) and NAA (1-naphthaleneacetic acid - 0.0; 0.3, or 0.5 mg L-1) in the development of adventitious shoots was evaluated for C. aurantium. Culture medium supplementation with BAP is not essential for the adventitious shoots development in the four genotypes studied when epicotyl segments-derived explants are used. In general, culture media supplementation with BAP decreased the percentage of responsive explants excepted for C. sinensis cv. 'Natal' and C. limonia when the concentrations of 1.5 and 2.0 mg/L were used. The presence of cytokinin, in concentrations up to 2 mg/L, stimulated the in vitro organogenesis when internodal segments-derived explants were used for C. limonia and C. aurantium. For C. aurantium no adventitious shoots developed in explants (internodal segments) cultured in basal culture medium, without BAP supplementation. Although no statistic differences could be detected, culture media supplementation with the combination of BAP and NAA favored the development of adventitious shoots in C. aurantium. The best concentration of NAA varied according to BAP concentration. The results presented herein, show that Citrus in vitro organogenesis depends on the interaction of culture medium composition, explant differentiation level, and genotype.

scholarly journals High Efficiency Direct Shoot Organogenesis from Leaf Segments ofAerva lanata(L.) Juss. Ex Schult by Using Thidiazuron

2014 ◽  
Vol 2014 ◽  
pp. 1-6 ◽  
Author(s):  
K. Varutharaju ◽  
C. Soundar Raju ◽  
C. Thilip ◽  
A. Aslam ◽  
A. Shajahan
Keyword(s):  
Shoot Organogenesis ◽  
Adventitious Shoots ◽  
Leaf Explants ◽  
Naphthaleneacetic Acid ◽  
Scale Production ◽  
Ms Medium ◽  
Leaf Segments ◽  
Direct Shoot Organogenesis ◽  
Direct Shoot

An efficient protocol for direct shoot organogenesis has been developed for the medicinal plantAerva lanata(L.) Juss. ex Schult. Regeneration was achieved from leaf segments of 20 days oldin vitroplantlets raised on Murashige and Skoog (MS) medium containing 0.25–2.0 mg L−1thiadiazuron (TDZ), 3% sucrose, and 0.8% agar. After 21 days of culture incubation, maximum number of shoot organogenesis (23.6 ± 0.16) was obtained on medium containing 1.0 mg L−1TDZ. The shoots were able to producein vitroflowers on medium containing 1.0 mg L−1TDZ in combination with 0.25–0.5 mg L−1  α-naphthaleneacetic acid (NAA). Histological observation showed that the epidermal cells of the leaf explants exhibited continuous cell division led to formation of numerous dome shaped meristematic protrusions and subsequently developed into adventitious shoots. Upon transfer of shootlets to half strength MS medium containing 1.0 mg L−1indole-3-butyric acid (IBA), around 86% of the regenerated shoots formed roots and plantlets. Rooted plants were hardened and successfully established in the soil at the survival rate of 92%. The regeneration protocol developed in this study provides an important method of micropropagation of this plant. Furthermore, this protocol may be used for a large scale production of its medicinally active compounds and genetic transformations for further improvement.

Microspore embryogenesis in anther culture of three species of Populus and regeneration of dihaploid plants of Populustrichocarpa

1993 ◽  
Vol 23 (9) ◽  
pp. 1821-1825 ◽  
Author(s):  
Snorri Baldursson ◽  
Peter Krogstrup ◽  
Jens Viktor Nørgaard ◽  
Sven Bode Andersen
Keyword(s):  
Chromosome Number ◽  
Growth Regulator ◽  
Woody Plant ◽  
Adventitious Shoots ◽  
Microspore Embryogenesis ◽  
Naphthaleneacetic Acid ◽  
Chromosome Counts ◽  
Low Frequencies ◽  
Woody Plant Medium

Microspore embryogenesis was induced from in vitro cultured anthers of Populusbalsamifera L., Populusmaximowiczii A. Henry, and Populustrichocarpa Torr. & Gray. Embryoids were formed at low frequencies on a modified Murashige and Skoog's medium, supplemented with 5 μM 6-benzylaminopurine, 5.1 mM L-glutamine, and 6% maltose. Growth regulator combinations (0–10 μM 6-benzylaminopurine and naphthaleneacetic acid) affected embryogenesis only slightly but formation of nonembryogenic callus from the anthers increased with increasing concentration of naphthaleneacetic acid. One donor clone of P. trichocarpa produced 54 embryoids from as many anthers during the two years of study. Adventitious shoots were obtained from 31 of these embryoids on woody plant medium with 2.5 μM 6-benzylaminopurine and 0.005 μM naphthaleneacetic acid. Adventitious shoots from 25 different embryoids were successfully rooted on woody plant medium containing 0.25 μM indole-3-butyric acid and transplanted to soil. Isozyme analysis confirmed microspore origin of all plants studied, and chromosome counts revealed that most of them had doubled their chromosome number spontaneously.

scholarly journals In vitro culture of Vriesea gigantea and Vriesea philippocoburgii: two vulnerable bromeliads native to Southern Brazil

2005 ◽  
Vol 48 (5) ◽  
pp. 717-722 ◽  
Author(s):  
Annette Droste ◽  
Anelise Machado da Silva ◽  
Adriana Vieira Matos ◽  
Júlia Winck de Almeida
Keyword(s):  
In Vitro Culture ◽  
Germination Rate ◽  
Adventitious Shoots ◽  
Basal Medium ◽  
Naphthaleneacetic Acid ◽  
Southern Brazil ◽  
Regeneration Medium ◽  
Best Response ◽  
Set Up

Micropropagation studies were carried out using the seeds for establishing an in vitro culture of Vriesea gigantea and Vriesea philippocoburgii. Germination rate of V. gigantea was higher than of V. philippocoburgii. Plantlets of V. philippocoburgii gave rise to many adventitious shoots when cultivated in Knudson basal medium. In contrast, for V. gigantea, a higher salts-concentration was needed, so that the number of shoots was increased by Murashige and Skoog medium. Addition of activated charcoal and naphthaleneacetic acid in regeneration medium allowed the growth of shoots and the formation of roots, confirming the success of in vitro culture. The differences in expression of the genotypes reinforce the need of more research in order to set up the conditions that could offer the best response of the specific tissues.

scholarly journals Shoot Regeneration Rates of Perennial Phlox are Dependant on Cultivar and Explant Type

HortScience ◽  
2004 ◽  
Vol 39 (6) ◽  
pp. 1373-1377 ◽  
Author(s):  
Margarita Fraga ◽  
Mertxe Alonso ◽  
Marisé Borja
Keyword(s):  
Adventitious Shoots ◽  
Leaf Explants ◽  
Naphthaleneacetic Acid ◽  
Meristem Culture ◽  
Ms Medium ◽  
Virus Elimination ◽  
Open Flower ◽  
Regeneration Rate ◽  
Shoot Initiation

Meristem culture and/or thermotherapy were used for virus elimination from ornamental Phlox paniculata L. (`Blue Boy', `Orange perfection' and `Starfire') mother plants. Shoot tip, leaf, node and flower ovary explants collected from greenhouse-maintained virus free plants were cultured in vitro for shoot initiation. Adventitious shoot initiation was observed on Murashige and Skoog (MS) medium containing the cytokinin BA with or without the auxin NAA. The addition of 0.4 mg·L-1 thiamine, 0.4 mg·L-1 folic acid, and 40 mg·L-1 adenine sulfate to the MS medium did not improve the regeneration rate. Multiplication and rooting were genotype dependent. Blue Boy and Orange Perfection cultivars regenerated the maximum number of shoots from leaf explants. `Blue Boy' leaf explants from in vitro plants had a lower regeneration rate than explants from greenhouse plants. Cultivar `Starfire' had the highest shoot formation with open flower ovary explants and failed to regenerate from leaf explants. In vitro rooting of adventitious shoots in the presence of auxins (IAA, NAA, or IBA) with or without BA was less effective than ex vitro rooting. Chemical names used: 6-benzyladenine (BA); indole-acetic acid (IAA); indole-3-butyric acid (IBA); α-naphthaleneacetic acid (NAA).

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