In vitro direct and indirect regeneration of promising lavandin cultivars

The medicinal plant, Aspilia africana, has been traditionally used in several African countries to treat many diseases such as tuberculosis, cough, inflammation, malaria, osteoporosis, and diabetes. In this study, we developed a protocol for in vitro propagation of A. africana using indirect shoot organogenesis from leaf and root explants of in vitro-grown seedlings and assessed the tissues at different developmental stages. The highest callus induction (91.9 ± 2.96%) from leaf explants was in the Murashige and Skoog (MS) medium augmented with 1.0 mg/L 6-Benzylaminopurine (BAP) and 1.0 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D) while from root explants, the highest callus induction (92.6 ± 2.80%) was in the same plant tissue culture medium augmented with 0.5 mg/L BAP and 1.0 mg/L 2,4-D. The best shoot regeneration capacity from leaf-derived calli (i.e., 80.0 ± 6.23% regeneration percentage and 12.0 ± 6.23 shoots per callus) was obtained in medium augmented with 1.0 mg/L BAP and 0.05 mg/L α-Naphthaleneacetic acid (NAA); the best regeneration capacity for root-derived calli (i.e., 86.7 ± 6.24% shoot regeneration percentage and 14.7 ± 1.11 shoots per callus) was obtained in the MS medium augmented with 1.0 mg/L BAP, 0.05 mg/L NAA, and 0.1 mg/L Thidiazuron (TDZ). Regenerated plantlets developed a robust root system in 1/2 MS medium augmented with 0.1 mg/L NAA and had a survival rate of 93.6% at acclimatization. The in vitro regenerated stem tissue was fully differentiated, while the young leaf tissue consisted of largely unorganized and poorly differentiated cells with large intercellular airspaces typical of in vitro leaf tissues. Our study established a protocol for the indirect regeneration of A. africana and offers a basis for its domestication, large-scale multiplication, and germplasm preservation. To the best of our knowledge, this is the first study to develop an indirect regeneration protocol for A. africana and conduct anatomical assessment through the different stages of development from callus to a fully developed plantlet.

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In vitro regeneration of pumpkin (cucurbita maxima) through shoot apical meristem

Journal of Bio-Science ◽

10.3329/jbs.v18i0.8784 ◽

1970 ◽

Vol 18 ◽

pp. 104-107 ◽

Cited By ~ 1

Author(s):

ME Haque ◽

D Rezwana ◽

MA Islam ◽

B Sikdar

Keyword(s):

Shoot Apical Meristem ◽

Apical Meristem ◽

In Vitro Regeneration ◽

Total Yield ◽

Meristem Culture ◽

Culture Vessel ◽

Cucurbita Maxima ◽

Indirect Regeneration

Context: Pumpkin (Cucurbita maxima) is very important vegetable crop. It is damaged by several types of diseases, especially viral diseases which reduce the total yield of pumpkin. This is very difficult problem and such type of problem can be overcome by meristem culture. Objectives: To develop suitable protocols for indirect regeneration of pumpkin plant through shoot apical meristem. Materials and Methods: Meristems were isolated from 15-21 days in vivo grown plants by collecting shoot tips and were prepared (0.2-0.5 mm) under 4X zoom stereo-microscope remaining two leaves primordia. Inoculation of explants was made singly per culture vessel in semisolid MS fortified with different concentrations and combinations of cytokinin and auxin. Results: Among different concentrations, 1.0 mg/l BAP showed best response for callus induction. Calli were sub cultured for shoot formation in MS containing BAP singly or combination with GA3 and 1.5 mg/l BAP + 0.1 mg/l GA3 gave better result. In vitro regenerated shoots were rooted well in ½ strength MS with 0.5 mg/l IBA and micro plants were acclimatized successfully in natural condition. Conclusion: Indirect regeneration of pumpkin plants through shoot apical meristem has been established. Keywords: Growth regulators; in vitro regeneration; Pumpkin; Cucurbita maxima. DOI: http://dx.doi.org/10.3329/jbs.v18i0.8784 JBS 2010; 18(0): 104-107

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Indirect Regeneration and Assessment of Genetic Fidelity of Acclimated Plantlets by SCoT, ISSR, and RAPD Markers in Rauwolfia tetraphylla L.: An Endangered Medicinal Plant

BioMed Research International ◽

10.1155/2019/3698742 ◽

2019 ◽

Vol 2019 ◽

pp. 1-14 ◽

Cited By ~ 15

Author(s):

Gulab Khan Rohela ◽

Phanikanth Jogam ◽

Prasad Bylla ◽

Christopher Reuben

Keyword(s):

Medicinal Plant ◽

Rapd Marker ◽

Genetic Fidelity ◽

Mother Plant ◽

Garden Soil ◽

Naphthalene Acetic Acid ◽

Stem Explants ◽

Clonal Multiplication ◽

Indirect Regeneration

Rauwolfia tetraphylla L. is an important medicinal plant species which is well known for its pharmaceutically important alkaloids. In the present study, we are reporting about its conservation by in vitro clonal multiplication through the standardized protocol of indirect regeneration by using leaf and stem based callus and assessment of genetic fidelity of acclimated plantlets by start codon targeted (SCoT), inter simple sequence repeats (ISSR), and randomly amplified polymorphic DNA (RAPD) marker based analysis. Initially friable callus was induced in maximum amounts (378.7, 323.8, and 412.8 in mg) from leaf, root, and stem explants on Murashige and Skoog (MS) media supplemented with 5.0 mg/L, 3.0 mg/L of 2,4-dichlorophenoxyacetic acid (2,4-D) and 5.0 mg/L of naphthalene acetic acid (NAA), respectively. Shoot regeneration with the maximum number of shoot buds (25 and 20) was obtained from leaf and stem calluses on MS media supplemented with TDZ (0.25 mg/L) + BAP (2 mg/L). The regenerated shoots were rooted successfully with maximum rooting percentage of 98.0 on full strength MS media amended with IAA (1.0 mg/L) and IBA (1.0 mg/L). The regenerated plantlets were hardened using 2:1 ratio of sterile garden soil and sand, followed by acclimatization in field conditions with 86% of survival. SCoT, ISSR, and RAPD primers based polymerase chain reaction (PCR) analysis was carried out to check possible genetic variations in micro propagated plants in comparison with mother plant. Among the ten SCoT (S), ISSR (R), and RAPD (OPA) primers used, S2, R10, and OPA3 has given good amplification with scorable DNA bands. The results revealed that the regenerated plants did not have any polymorphism with mother plant. Hence, the in vitro regenerated R. tetraphylla plantlets were confirmed as true-to-type.

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Indirect regeneration of Withania somnifera and comparative analysis of withanolides in in vitro and greenhouse grown plants

Biologia Plantarum ◽

10.1007/s10535-010-0063-6 ◽

2010 ◽

Vol 54 (2) ◽

pp. 357-360 ◽

Cited By ~ 25

Author(s):

Y. H. Dewir ◽

D. Chakrabarty ◽

S. -H. Lee ◽

E. -J. Hahn ◽

K. -Y. Paek

Keyword(s):

Comparative Analysis ◽

Withania Somnifera ◽

Indirect Regeneration

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The effects of genotypes and media composition on callogenesis, regeneration and cell suspension culture of chamomile (Matricaria chamomilla L.)

PeerJ ◽

10.7717/peerj.11464 ◽

2021 ◽

Vol 9 ◽

pp. e11464

Author(s):

Aqeel Ahmad ◽

Muhammad Tahir ul Qamar ◽

Almeera Shoukat ◽

Mehtab Muhammad Aslam ◽

Mohsin Tariq ◽

...

Keyword(s):

Suspension Culture ◽

Cell Suspension ◽

Cell Suspension Culture ◽

In Vitro Regeneration ◽

Sustainable Production ◽

Conservation Strategy ◽

Direct Regeneration ◽

Ms Medium ◽

Indirect Regeneration

Background Chamomile is an important herb being used widely for medicinal purposes. Its multitherapeutic, cosmetic, and nutritional values have been established through years of traditional and scientific use and research. Increased use of medicinal plants necessitates rational use as well as sustainable production of such genetic resources. Plant in vitro micro-propagation poses unique opportunities for sustainable production of medicinal herbs, their regrowth and conservation. The present study aimed to investigate the effects of different explants, plant growth regulators (PGRs) combinations and media type on callogenesis, in vitro regeneration and cell suspension of six chamomile genotypes to enhance its sustainable production. Methods The shoot, lateral sprout, and leaf derived explants of six chamomile genotypes including Isfahan, Shiraz, Kazeron, Goral, Sharokashari and Presso were used for direct and indirect regeneration. For indirect regeneration various doses of NAA and kinetin were used to induce calli which were cultured on MS media containing PGRs for direct and indirect regeneration. Later, cell suspension was established and morphological characterization of CrO3 stained cells was carried out using microscopy. Results and Discussion Our findings revealed that the highest callus percentage and callus volume were observed from lateral sprouts and shoots of genotype Isfahan on MS medium containing 1 mg/L NAA and 1 mg/L kinetin. The in vitro regeneration was found to be genotype dependent while 77% and 77.5% was the highest percentage for indirect and direct regeneration, respectively. Additionally, the maximum shoot number (two shoots/explant) and shoot length (2.22 cm) were also observed in Isfahan genotype. Cell suspension culture showed the highest fresh weight (18.59 g) and dry weight (1.707 g) with 0.75 g inoculum of the callus derived from lateral sprouts cultured on MS medium. Microscopy of CrO3 stained cells was carried on each 3rd day for 27 days that revealed larger and spongier cells in the early days as compared to final days when the cell number was greater but cell size was smaller. Conclusion The callogenesis, organogenesis, and cell suspension culture of chamomile may be genotype dependent. Hence, optimization of media ingredients and culture conditions is of utmost importance for devising tissue culture based conservation strategy of any chamomile genotype and secondary metabolite production.

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