scholarly journals Genotype-dependent mass somatic embryogenesis: a chance to recover extinct populations of Pulsatilla vulgaris Mill.

2021 ◽  
Author(s):  
Justyna Żabicka ◽  
Piotr Żabicki ◽  
Aneta Słomka ◽  
Monika Jędrzejczyk-Korycińska ◽  
Teresa Nowak ◽  
...  
Keyword(s):  
Somatic Embryogenesis ◽  
Critical Point ◽  
Adventitious Shoots ◽  
Genetic Fidelity ◽  
Issr Markers ◽  
Molecular Techniques ◽  
Ex Situ ◽  
Experimental Conditions ◽  
Regenerated Plantlets

Abstract The paper presents a technique for micropropagation of endangered in Europe and extinct in Poland Pulsatilla vulgaris for ex situ conservation of the genetic resources. Genotype-dependent induction of somatic embryogenesis and rooting was revealed in series of two experiments (I and II) performed under the same experimental conditions. Shoot tips of seedlings were the best explants in both experiments and Murashige and Skoog (MS) medium supplemented with 0.25 or 0.5 mg L−1 BAP was suitable for induction of somatic embryos (SE) and adventitious shoots. Mass SE was obtained in experiment I after explants transfer on ½ MS (2% sucrose) + 0.45 mg L−1 B1 and extending culture to 2–3 months without passages. Rooting of adventitious shoots was a critical point. Out of seven rooting media used in experiment I, only two, ½ MS hormone free (2% sucrose) + 0.45 mg L−1 B1 or MS + 5 mg L−1 NAA + 3.76 mg L−1 B2 resulted in altogether 36.4% rooted shoots. In experiment II, somatic embryogenesis, rooting and acclimatization of adventitious shoots failed. Regenerated plantlets and seedlings converted from SE from experiment I were acclimatized to ex vitro conditions. Both genome size, determined by flow cytometry, and genetic diversity analyzed by ISSR markers, confirmed the compatibility of regenerants from experiment I with P. vulgaris initial seedlings and commercial cultivar. Regenerants obtained in experiment II differed genetically from the regenerants of experiment I and cultivar. Propagated in vitro tissues/organs (SE, adventitious shoots) of P. vulgaris could be a source of material for cryopreservation, artificial seed production and/or for acclimatization of regenerated plantlets and could be used for restoration of the extinct populations. Key Message The micropropagation technique via organogenesis and somatic embryogenesis of endangered in Europe pasqueflower was developed as a tool for species recovery. The critical point is that somatic embryogenesis is genotype-dependent, which affects the repeatability of the experiments and also imposes applying molecular techniques to confirm the genetic fidelity of the regenerants with the initial material and other genotypes.

scholarly journals Metabolic profiling, in vitro propagation, and genetic assessment of the endangered rare plant Anarrhinum pubescens

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Asmaa Abdelsalam ◽  
Ehab Mahran ◽  
Kamal Chowdhury ◽  
Arezue Boroujerdi
Keyword(s):  
In Vitro Propagation ◽  
Genetic Fidelity ◽  
Ex Situ ◽  
Wild Plant ◽  
Rare Plant ◽  
Chemical Profiling ◽  
Nodal Cutting ◽  
Regenerated Plants ◽  
Plant Shoots

Abstract Background Anarrhinum pubescens Fresen. (Plantaginaceae) is a rare plant, endemic to the Saint Catherine area, of South Sinai, Egypt. Earlier studies have reported the isolation of cytotoxic and anti-cholinesterase iridoid glucosides from the aerial parts of the plant. The present study aimed to investigate the chemical profiling of the wild plant shoots as well as establish efficient protocols for in vitro plant regeneration and proliferation with further assessment of the genetic stability of the in vitro regenerated plants. Results Twenty-seven metabolites have been identified in wild plant shoots using the Nuclear Magnetic Resonance (NMR) spectroscopy. The metabolites include alkaloids, amino acids, carbohydrates, organic acids, vitamins, and a phenol. In vitro propagation of the plant was carried out through nodal cutting-micropropagation and leaf segment-direct organogenesis. The best results were obtained when nodal cutting explants were cultured on Murashige and Skoog medium with Gamborg B5 vitamins supplemented with 6-benzylaminopurine (BAP) (1.0 mg/L) and naphthaleneacetic acid (NAA) (0.05 mg/L), which gave a shoot formation capacity of 100% and a mean number of shoots of 27.67 ± 1.4/explant. These shoots were successfully rooted and transferred to the greenhouse and the survival rate was 75%. Genetic fidelity evaluation of the micropropagated clones was carried out using random amplified polymorphic DNA (RAPD) and inter simple sequence repeat (ISSR) molecular markers. Jaccard’s similarity coefficient indicated a similarity as high as 98% and 95% from RAPD and ISSR markers, respectively. Conclusions This study provides the chemical profiling of the aerial part of Anarrhinum pubescens. Moreover, in vitro regeneration through different tissue culture techniques has been established for mass propagation of the plant, and the genetic fidelity of the in vitro regenerated plants was confirmed as well. Our work on the in vitro propagation of A. pubescens will be helpful in ex situ conservation and identification of bioactive metabolites.

scholarly journals Silver nanoparticles induce genetic, biochemical, and phenotype variation in chrysanthemum

2020 ◽  
Vol 143 (2) ◽  
pp. 331-344
Author(s):  
Alicja Tymoszuk ◽  
Dariusz Kulus
Keyword(s):  
Silver Nanoparticles ◽  
In Vitro Regeneration ◽  
Adventitious Shoots ◽  
Issr Markers ◽  
Phenotype Variation ◽  
Inter Simple Sequence Repeats ◽  
Tremendous Progress ◽  
Genetic Level ◽  
User Friendly

Abstract Despite the tremendous progress in breeding, novel and user-friendly techniques of plant improvement are desirable. The study aimed to analyze the usefulness of silver nanoparticles (AgNPs) in the breeding of chrysanthemum: one of the top ornamental plant species. In vitro regeneration of adventitious shoots from internodes of chrysanthemum ‘Lilac Wonder’ was induced on the modified Murashige and Skoog (MS) medium supplemented with 0.6 mg L−1 6-benzylaminopurine (BAP), 2 mg L−1 indole-3-acetic acid (IAA) and AgNPs at 0, 5, 10 and 20 ppm concentration. The efficiency of callogenesis and caulogenesis were analyzed after 10 weeks of culture. The concentration of chlorophylls, carotenoids, and phenolic compounds in shoots and calli were estimated. Plants obtained from 20 ppm AgNPs treatment were additionally analyzed on the genetic level using randomly amplified polymorphic DNA (RAPD) and inter simple sequence repeats (ISSR) markers. In vitro rooted shoots were acclimatized in the glasshouse and subjected to biochemical and phenotype stability evaluation. AgNPs at the highest concentration (20 ppm) suppressed both callogenesis and caulogenesis in vitro. The concentration of metabolites in callus was stable, regardless of AgNPs treatment, except for carotenoids which production was enhanced by 20 ppm AgNPs. In contrast, the content of chlorophyll a and b in shoots varied depending on AgNPs treatment. Polymorphic loci were detected in 12 and 9 AgNPs-treated-plants by RAPD and ISSR markers, respectively (one of which was common to both marker systems). Rooting and acclimatization were fully successful in all experimental combinations. Phenotype alternations were detected in six plants; one from 10 ppm AgNPs treatment and five from 20 ppm treatment. They included variation in pigment content (anthocyanins and carotenoids) and/or inflorescence shape. Interestingly, only two plants revealed both genetic and phenotype polymorphisms. No genetic or phenotype variation was detected in the control plants. In conclusion, AgNPs can be used in chrysanthemum breeding.

scholarly journals Nutrient optimization for improved in vitro plant regeneration in Eclipta alba (L.) Hassk. and assessment of genetic fidelity using RAPD analysis

2015 ◽  
Vol 24 (2) ◽  
pp. 223-234
Author(s):  
Shruti Bardar ◽  
Varsha Khurana Kaul ◽  
Sumita Kachhwaha ◽  
SL Kothari
Keyword(s):  
Basal Medium ◽  
Genetic Fidelity ◽  
Ex Situ ◽  
Nutrient Level ◽  
Induction Medium ◽  
Eclipta Alba ◽  
Shoot Bud ◽  
Bud Induction ◽  
Regeneration Response

This study highlights the effect of different inorganic micronutrients like copper, cobalt, molybdenum, zinc, boron, iodine, iron and manganese in accelerating and amplifying in vitro shoot bud induction and proliferation of a medicinally important plant, Eclipta alba (L.) Hassk. Direct shoot bud induction was observed on MS fortified with Kn (2 mg/l). However, maximum number of shoots was achieved when GA3, 0.5 mg/l was added to induction medium along with 1?M copper sulphate (ten times the normal MS level). Optimization of nutrient level in the basal medium promoted maximum regeneration response from both shoot tips and nodal explants. Elongated shoots were rooted in MS supplemented with IBA, 1.0 mg/l. Healthy, green plantlets with well developed roots, flowered normally in the field. Genetic stability of micropropagated plantlets was evaluated using RAPD markers. The amplification products were monomorphic in micropropagated plantlets and similar to those of mother plant revealing the genetic uniformity of plantlets. The regeneration protocol is highly efficient and reproducible so would be useful for mass multiplication, ex situ conservation and genetic transformation of E. alba (L.) Hassk.Plant Tissue Cult. & Biotech. 24(2): 223-234, 2014 (December)

scholarly journals Micropropagation and Genetic Fidelity of the Regenerants of Aglaonema ‘Valentine’ Using Randomly Amplified Polymorphic DNA

HortScience ◽  
2016 ◽  
Vol 51 (4) ◽  
pp. 398-402 ◽  
Author(s):  
Mohammed Elsayed El-Mahrouk ◽  
Yaser Hassan Dewir ◽  
Yougasphree Naidoo
Keyword(s):  
In Vitro Regeneration ◽  
Shoot Proliferation ◽  
Genetic Fidelity ◽  
Naphthalene Acetic Acid ◽  
Axillary Shoot ◽  
Ms Medium ◽  
Randomly Amplified Polymorphic Dna ◽  
Simple Protocol ◽  
Regenerated Plantlets

The present study reports a simple protocol for in vitro regeneration of Aglaonema ‘Valentine’ using axillary shoot explants for rapid multiplication and production of true-to-type plants. Different concentrations of benzyladenine (BA; 0, 1, 3, 5, and 7 mg·L−1), kinetin (Kin; 0, 1, 3, 5, and 7 mg·L−1), thidiazuron (TDZ; 0, 0.5, 1.0, 1.5, and 2.0 mg·L−1), naphthalene acetic acid (NAA; 0, 0.5, and 1.0 mg·L−1), and indole-3-butyric acid (IBA; 0, 0.5, and 1.0 mg·L−1) were used for shoot regeneration. The highest shoot proliferation (5.0) was obtained on Murashige and Skoog (MS) medium supplemented with 1.5 mg·L−1 TDZ and 1 mg·L−1 NAA. In vitro rooting was easily achieved with 100% at all concentrations of NAA and IBA supplemented to half- or full-strength MS medium. Regenerated plantlets were acclimatized in greenhouse with 100% survival rate. Randomly amplified polymorphic DNA (RAPD) analysis confirmed the genetic fidelity of the regenerated plantlets and mother plant.

scholarly journals In vitro morphogenetic responses from obligatory apomictic Taraxacum belorussicum Val. N. Tikhom seedlings explants

2019 ◽  
Vol 139 (3) ◽  
pp. 505-522 ◽  
Author(s):  
Adrianna Gałuszka ◽  
Maciej Gustab ◽  
Monika Tuleja
Keyword(s):  
Somatic Embryogenesis ◽  
Plant Material ◽  
Molecular Mechanisms ◽  
Adventitious Shoots ◽  
Indirect Organogenesis ◽  
Regenerated Plants ◽  
Blue Solution ◽  
Meristem Explants ◽  
Scanning Electron

Abstract Taraxacum belorussicum Val. N. Tikhom, a poorly known and obligatory apomictic species, is an attractive plant material for studying the embryological, genetic and molecular mechanisms of apomixis. This work aims to obtain an efficient protocol for Taraxacum belorussicum regeneration. Four types of explants (cotyledons, hypocotyls, meristems and roots) that were taken from 2-weeks-old seedlings were used for in vitro cultures, and a fast and efficient protocol of T. belorussicum regeneration was obtained. Various ½ MS-based media containing IAA (5.71 µM), TDZ (4.54 µM) and PSK (100 nM) were chosen to assess the morphogenetic abilities of selected T. belorussicum explants. Studies on the role of PSK were done in three independent experiments, where the most significant factors were always light and darkness. All explants produced callus by the third day of culture and adventitious shoots after 7 days, although in an asynchronous indirect manner, and with different intensities for all explant types. The most preferred medium culture for hypocotyl, cotyledon and meristem explants was ½ MS + TDZ, and ½ MS + IAA + TDZ + PSK for roots which were the only explant sensitive to PSK. A short darkness pretreatment (8 days) in PSK medium was found suitable to enhance organogenesis. Secondary organogenesis was observed for regenerated plants on meristem explants from the ½ MS + IAA + TDZ + PSK medium. A weak somatic embryogenesis was observed for hypocotyl and cotyledon explants from ½ MS + IAA + TDZ and ½ MS + IAA + TDZ + PSK media. Histological and scanning electron microscope images (SEM) of T. belorussicum confirmed indirect organogenesis and somatic embryogenesis. Plant material treated with aniline blue solution revealed the presence of callose in the cell walls of cotyledon and hypocotyl explants. The presence of extracellular matrix (ECM) and heterogenic structure of callus was also verified by scanning electron microscopy and light microscopy, confirming the high morphogenetic ability of T. belorussicum.

scholarly journals The Effect of Different Factors upon the in vitro Propagation in Quercus robur and Q. frainetto by Somatic Embryogenesis

2011 ◽  
Vol 39 (1) ◽  
pp. 288 ◽  
Author(s):  
Adrian Ioan TIMOFTE ◽  
Doru PAMFIL ◽  
Magdalena PALADA-NICOLAU ◽  
Claudia Simona TIMOFTE
Keyword(s):  
Somatic Embryogenesis ◽  
Significant Influence ◽  
Quercus Robur ◽  
Culture Medium ◽  
Clonal Propagation ◽  
Germination Percentage ◽  
Development Stage ◽  
Ex Situ ◽  
Conservation Of Genetic Resources

The somatic embryogenesis is an advanced method for clonal propagation and a useful tool for ex situ conservation of genetic resources. In this paper, the results of an experiment to investigate the influence of development stage of explants and culture medium on the germination percentage in two oak species (three provenances of Quercus robur and two provenances of Q. frainetto), are presented. A high significant influence of the development stage of explants and a significant influence of the interaction provenance x stage on the germination percentage were recorded for Q. robur explants, whilst no significant differences between the germination percentages against the nutritive media used were fould for both oak species.

scholarly journals Ex situ preservation in medium-term culture of the threathened taxon Dianthus nardiformis Janka

2021 ◽  
Vol 26 (2) ◽  
pp. 2416-2422
Author(s):  
IRINA HOLOBIUC ◽  
◽  
RODICA CATANĂ ◽  
FLORENȚA HELEPCIUC ◽  
CARMEN MAXIMILIAN ◽  
...  
Keyword(s):  
Somatic Embryogenesis ◽  
Ex Situ ◽  
In Vitro Growth ◽  
Growth Reduction ◽  
Medium Term ◽  
Time Intervals ◽  
Qualitative And Quantitative ◽  
Unlimited Time ◽  
In Vitro Response

Our aim was to elaborate an efficient and reproducible protocol for medium-term culture of the threatened taxon Dianthus nardiformis. To reduce the growth, sucrose, mannitol, polyethylene glycol, Abscisic acid and Jasmonic acid were tested. For assessing the in vitro response, the growth and regeneration were registered after different time intervals. Mannitol is the most effective for medium-term preservation viable cultures which can be maintained unlimited time through transfer at every 3 months. In its presence, somatic embryogenesis was induced and in vitro growth in the minimal cultures was reduced between 9 and 12 times comparing to the control. Antioxidant enzymes assay revealed qualitative and quantitative differences among the experimental variants, and also between different concentrations of the same compound in correlation with the growth reduction and regeneration. POX was the most suitable to detect the efficiency of different treatments to induce medium-term cultures.

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