Development of an efficient protocol for high frequency in vitro regeneration of a horticultural plant Primulina tamiana (B.L. Burtt) Mich. Möller & A. Webber

2014 ◽  
Vol 94 (7) ◽  
pp. 1281-1287 ◽  
Author(s):  
Priya Padmanabhan ◽  
Susan J. Murch ◽  
J. Alan Sullivan ◽  
Praveen K. Saxena
Keyword(s):  
Shoot Regeneration ◽  
High Frequency ◽  
De Novo ◽  
Shoot Organogenesis ◽  
In Vitro Regeneration ◽  
Leaf Explants ◽  
Root Induction ◽  
Horticultural Plant ◽  
Leaf Segments

Padmanabhan, P., Murch, S. J., Sullivan, J. A. and Saxena, P. K. 2014. Development of an efficient protocol for high frequency in vitro regeneration of a horticultural plant Primulina tamiana (B.L. Burtt) Mich. Möller & A. Webber. Can. J. Plant Sci. 94: 1281–1287. A rapid and efficient in vitro regeneration system was established for Primulina tamiana, an attractive ornamental plant of horticultural importance. The successful regeneration protocol employed induction of shoot organogenesis on leaf explants. Among the various plant growth regulators evaluated, thidiazuron (TDZ) proved to be the most effective in inducing rapid de novo shoot formation. The highest shoot regeneration frequency within the shortest time from leaf explants was obtained on medium enriched with 2.5 µM TDZ. Furthermore, leaf segments were found to be a more suitable explant for in vitro shoot regeneration compared with petiole explants. The abaxial side of the leaf segments, which was in contact with the culture medium, generated more shoots than the adaxial side. Scanning electron microscopic studies carried out at various stages of de novo shoot differentiation demonstrated that shoot organogenesis involved the formation of actively dividing regions from the epidermal and subepidermal layers of the explants. In vitro rooting of the regenerated shoots was attempted using two different auxins, IAA (3-indoleacetic acid) and 3-indolebutyric acid (IBA) (0, 0.1, 0.5, 1.0, 2.0, and 5.0 µM). The optimum concentration for root induction was found to be 5 µM IBA. Approximately, 98% of the regenerated plants survived transfer and acclimation to greenhouse conditions.

scholarly journals The effects of β-lactam antibiotics and hygromycin B on de novo shoot organogenesis in apple cv. Golden Delicious

2018 ◽  
Vol 70 (1) ◽  
pp. 179-190 ◽  
Author(s):  
Mariana Stanisic ◽  
Slavica Ninkovic ◽  
Jelena Savic ◽  
Tatjana Cosic ◽  
Nevena Mitic
Keyword(s):  
Shoot Regeneration ◽  
De Novo ◽  
Shoot Organogenesis ◽  
In Vitro Regeneration ◽  
Leaf Explants ◽  
Selection Procedure ◽  
Transformation Process ◽  
Hygromycin B ◽  
Golden Delicious

Since the genetic transformation of the apple is strongly genotype-dependent and generally inefficient, the evaluation of factors affecting shoot regeneration are crucial for the establishment of a successful transformation process. In this report, we evaluated the effects of the ?-lactam antibiotics meropenem and timentin on in vitro regeneration via de novo shoot organogenesis from leaf explants of apple cv. Golden Delicious, as well as on the growth of the Agrobacterium tumefaciens strain EHA 105, and compared them with the commonly used ?-lactam cefotaxime. Also, we report for the first time the effect of hygromycin B as a selective agent in the domesticated apple, as regards shoot regeneration and shoot multiplication efficiency. We observed that cefotaxime and timentin at concentrations higher than 100 mg L-1 were sufficient to prevent Agrobacterium growth during a two-week period, while meropenem exhibited an inhibitory effect on bacterial growth at all tested concentrations (25-150 mg L-1). Cefotaxime at a concentration of 300 mg L-1 increased the number of regenerated shoots per explant (9.39) in comparison with the control (7.67). In contrast to cefotaxime, meropenem and timentin caused a decrease in shoot regeneration efficiency, but larger and more developed shoots were obtained on meropenem (25-125 mg L-1) after the same period of cultivation. Hygromycin B at a concentration of 5 mg L-1 or higher completely inhibited shoot regeneration and induced explant tissue necrosis. Therefore, the selection procedure with a final concentration of 4 mg L-1 throughout organogenesis and 10 mg L-1 for further shoot growth and multiplication is recommended for an efficient transformation process in apple cv. Golden Delicious.

scholarly journals Integrating the Roles for Cytokinin and Auxin in De Novo Shoot Organogenesis: From Hormone Uptake to Signaling Outputs

2021 ◽  
Vol 22 (16) ◽  
pp. 8554
Author(s):  
Martin Raspor ◽  
Václav Motyka ◽  
Abdul Rasheed Kaleri ◽  
Slavica Ninković ◽  
Ljiljana Tubić ◽  
...  
Keyword(s):  
Shoot Regeneration ◽  
De Novo ◽  
Shoot Organogenesis ◽  
In Vitro Regeneration ◽  
Genetic Regulation ◽  
Auxin Signaling ◽  
Plant Tissues ◽  
Cultivated Plant ◽  
The Media

De novo shoot organogenesis (DNSO) is a procedure commonly used for the in vitro regeneration of shoots from a variety of plant tissues. Shoot regeneration occurs on nutrient media supplemented with the plant hormones cytokinin (CK) and auxin, which play essential roles in this process, and genes involved in their signaling cascades act as master regulators of the different phases of shoot regeneration. In the last 20 years, the genetic regulation of DNSO has been characterized in detail. However, as of today, the CK and auxin signaling events associated with shoot regeneration are often interpreted as a consequence of these hormones simply being present in the regeneration media, whereas the roles for their prior uptake and transport into the cultivated plant tissues are generally overlooked. Additionally, sucrose, commonly added to the regeneration media as a carbon source, plays a signaling role and has been recently shown to interact with CK and auxin and to affect the efficiency of shoot regeneration. In this review, we provide an integrative interpretation of the roles for CK and auxin in the process of DNSO, adding emphasis on their uptake from the regeneration media and their interaction with sucrose present in the media to their complex signaling outputs that mediate shoot regeneration.

scholarly journals Natural Variation in Plant Pluripotency and Regeneration

Plants ◽  
2020 ◽  
Vol 9 (10) ◽  
pp. 1261
Author(s):  
Robin Lardon ◽  
Danny Geelen
Keyword(s):  
De Novo ◽  
Shoot Organogenesis ◽  
In Vitro Regeneration ◽  
Relative Role ◽  
Root Explants ◽  
Experimental Systems ◽  
Complex Process ◽  
Insight Into ◽  
Molecular Framework

Plant regeneration is essential for survival upon wounding and is, hence, considered to be a strong natural selective trait. The capacity of plant tissues to regenerate in vitro, however, varies substantially between and within species and depends on the applied incubation conditions. Insight into the genetic factors underlying this variation may help to improve numerous biotechnological applications that exploit in vitro regeneration. Here, we review the state of the art on the molecular framework of de novo shoot organogenesis from root explants in Arabidopsis, which is a complex process controlled by multiple quantitative trait loci of various effect sizes. Two types of factors are distinguished that contribute to natural regenerative variation: master regulators that are conserved in all experimental systems (e.g., WUSCHEL and related homeobox genes) and conditional regulators whose relative role depends on the explant and the incubation settings. We further elaborate on epigenetic variation and protocol variables that likely contribute to differential explant responsivity within species and conclude that in vitro shoot organogenesis occurs at the intersection between (epi) genetics, endogenous hormone levels, and environmental influences.

scholarly journals In vitro regeneration protocol of Rauvolfia serpentina L.

2018 ◽  
Vol 53 (2) ◽  
pp. 133-138 ◽  
Author(s):  
S Khan ◽  
TA Banu ◽  
S Akter ◽  
B Goswami ◽  
M Islam ◽  
...  
Keyword(s):  
In Vitro Regeneration ◽  
Leaf Explants ◽  
Multiple Shoot ◽  
Nodal Segments ◽  
Root Induction ◽  
Ms Medium ◽  
Regeneration System ◽  
Rauvolfia Serpentina ◽  
Number Of Shoots

An efficient in vitro regeneration system was developed for Rauvolfia serpentina L. through direct and indirect organogenesis from nodal and leaf explants. Among the different growth regulators, MS medium supplemented with 2.0 mg/l BAP, 0.5mg/l IAA and 0.02mg/l NAA found best for the multiple shoot formation from nodal segments. In this combination 98% explants produced multiple shoots and the average number of shoots per explants is 13∙4. The frequency of callus induction and multiple shoot induction from leaves was highest 88% in MS medium supplemented with 2.0 mg/l BAP, where mean number of shoots/explants was 12.5. The highest frequency of root induction (80%) and mean number of roots/plantlets (10) were obtained on half strength of MS medium containing 0.2 mg/l IBA. The rooted plantlets were transferred for hardening following acclimatization and finally were successfully established in the field.Bangladesh J. Sci. Ind. Res.53(2), 133-138, 2018

scholarly journals In vitro Regeneration and Agrobacterium-mediated Genetic Transformation of Tomato (Lycopersicon esculentum Mill.)

1970 ◽  
Vol 19 (1) ◽  
pp. 101-111 ◽  
Author(s):  
Rakha Hari Sarker ◽  
Khaleda Islam ◽  
M.I. Hoque
Keyword(s):  
Lycopersicon Esculentum ◽  
Genetic Transformation ◽  
In Vitro Regeneration ◽  
Leaf Explants ◽  
Multiple Shoots ◽  
Multiple Shoot ◽  
Root Induction ◽  
Cotyledonary Leaf ◽  
Lycopersicon Esculentum Mill

Agrobacterium-mediated genetic transformation system has been developed for two tomato (Lycopersicon esculentum Mill.) varieties, namely Pusa Ruby (PR) and BARI Tomato-3 (BT-3). Prior to the establishment of transformation protocol cotyledonary leaf explants from the two varieties were cultured to obtain genotype independent in vitro regeneration. Healthy multiple shoot regeneration was obtained from the cut ends of cotyledonary leaf segments for both the varieties on MS containing 1.0 mg/l BAP and 0.1 mg/l IAA. The maximum root induction from the regenerated shoots was achieved on half the strength of MS medium supplemented with 0.2 mg/l IAA. The in vitro grown plantlets were successfully transplanted into soil where they flowered and produced fruits identical to those developed by control plants. Transformation ability of cotyledonary leaf explants was tested with Agrobacterium tumefaciens strain LBA4404 harboring binary plasmid pBI121, containing GUS and npt II genes. Transformed cotyledonary leaf explants were found to produce multiple shoots on MS containing 1.0 mg/l BAP and 0.1 mg/l IAA. Selection of the transformed shoots was carried out by gradually increasing the concentration of kanamycin to 200 mg/l since kanamycin resistant gene was used for transformation experiments. Shoots that survived under selection pressure were subjected to rooting. Transformed rooted plantlets were transferred to soil. Stable expression of GUS gene was detected in the various tissues from putatively transformed plantlets using GUS histochemical assay.  Key words: In vitro regeneration, transformation, tomato D.O.I. 10.3329/ptcb.v19i1.5004 Plant Tissue Cult. & Biotech. 19(1): 101-111, 2009 (June)

scholarly journals In Vitro Adventitious Shoot Regeneration through Direct and Indirect Organogenesis from Seedling-derived Hypocotyl Segments of Ficus religiosa L.: An Important Medicinal Plant

HortScience ◽  
2018 ◽  
Vol 53 (1) ◽  
pp. 55-61 ◽  
Author(s):  
Mohsen Hesami ◽  
Mohammad Hosein Daneshvar
Keyword(s):  
Shoot Regeneration ◽  
Shoot Organogenesis ◽  
Type I ◽  
Root Induction ◽  
Ms Medium ◽  
Regeneration Frequency ◽  
Indirect Organogenesis ◽  
Ficus Religiosa ◽  
Shoot Number

Ficus religiosa is an important industrial, medicinal, and ornamental plant, so in vitro regeneration is of high paramount in this valuable germplasm. Two efficient protocols were developed for indirect and direct shoot organogenesis through hypocotyl explants. In the first experiment, different concentrations of 2,4-dichlorophenoxyacetic acid (2,4-D) and indole butyric acid (IBA) (0.5, 1.0, and 1.5 mg·L−1) in combination with 6-benzyl amino purine (BAP) (ratio 10:1, respectively) were used for callus formation. Two types of callus were obtained from different concentrations of plant growth regulators (PGRs). Also, 2,4-D produced yellow-brownish and friable callus (Type I), whereas the green and compact callus (Type II) was achieved in IBA. The highest callus fresh weight (2.43 g) was observed in Murashige and Skoog (MS) medium containing 0.5 mg·L−1 2,4-D plus 0.05 mg·L−1 BAP. In the later experiments, various concentrations of thidiazuron (TDZ), 6-furfuryl amino purine (KN), and BAP (0.25, 0.5, 1.0, and 1.5 mg·L−1) in combination with IBA (ratio 10:1, respectively) were applied for shoot regeneration (direct and indirect organogenesis). In shoot regeneration from callus, the highest regeneration frequency (86.66%) and shoot number per callus (4.13) were achieved in MS medium supplemented with 1.5 mg·L−1 BAP plus 0.15 mg·L−1 IBA from type I calli. However, no regeneration was observed in type II calli. In direct shoot regeneration, the highest regeneration frequency (96.66%) and shoot number (6.26) were obtained in the medium mentioned previously. In root induction experiment, different concentrations of naphthalene acetic acid (NAA) and IBA alone or in combination were applied, and MS medium containing 2.0 mg·L−1 IBA along with 0.1 mg·L−1 NAA was the best hormonal balance for root induction. The rooted plantlets’ survival rate was more than 95% in the acclimatization stage. These results demonstrated that the direct regeneration method provides more shoot regeneration frequency and take a less time for shoot organogenesis than the indirect regeneration method. Based on our knowledge, this study is the first report of direct and indirect shoot organogenesis of F. religiosa via hypocotyl from in vitro–grown seedling.

scholarly journals High Frequency In vitro Regeneration of Gynura procumbens (Lour.) Merr.

2017 ◽  
Vol 27 (2) ◽  
pp. 207-216
Author(s):  
Tanjina Akhtar Banu ◽  
Barna Goswami ◽  
Shahina Akter ◽  
Mousona Islam ◽  
Tammana Tanjin ◽  
...  
Keyword(s):  
In Vitro Regeneration ◽  
Leaf Explants ◽  
Multiple Shoots ◽  
Multiple Shoot ◽  
Nodal Segments ◽  
Nodal Segment ◽  
Root Induction ◽  
Petiole Explants ◽  
Number Of Shoots

An efficient rapid in vitro regeneration protocol was described from nodal segment, leaf and petiole explants. MS medium supplemented with 1.0 mg/l BAP and 0.5 mg/l IAA was found best for the multiple shoot formation from nodal segments. In this combination 99% explants produced multiple shoots and the average number of shoots per explants was 20.1 ± 1.96. For petiole and leaf explants best response was observed on MS supplemented with 2.0 mg/l BAP, 1 mg/l IAA and 0.5 mg/l Kn. Petiole explants produced highest mean number of shoots/explant (22.9 ± 1.728) among the three explants when the explants were cultured on MS with 2.0 mg/l BAP, 1 mg/l IAA and 0.5 mg/l Kn. The highest frequency of root induction (100%) and mean number of roots/plantlets (11.75) were obtained on MS. The rooted plantlets were transferred for hardening following acclimatization and finally were successfully established in the field.Plant Tissue Cult. & Biotech. 27(2): 207-216, 2017 (December)

scholarly journals IN VITRO Regeneration of Bina Mungbean Varieties

2015 ◽  
Vol 7 (2) ◽  
pp. 47-52 ◽  
Author(s):  
S Mojumder ◽  
MD Hossain ◽  
MS Haque ◽  
KM Nasiruddin
Keyword(s):  
Shoot Regeneration ◽  
Callus Induction ◽  
Root Length ◽  
In Vitro Regeneration ◽  
Root Induction ◽  
Ms Medium ◽  
Root Initiation ◽  
Root Proliferation ◽  
Callus Initiation

The experiment was conducted to develop an efficient protocol for in vitro regeneration of mungbean (Vignaradiata) on the aspect of regeneration potentiality of two mungbean varieties (BINA mung 5 and BINA mung 7) as influenced by different combinations of growth regulators supplemented with MS medium. Cotyledon explant of both varieties was used for the present study. Data were collected for various characters of callus initiation, shoot regeneration and root proliferation. Initiation of callus (%) and required days for its initiation and weight of callus were influenced significantly due to the effect of varieties where BINA mung 5 produced more callus induction (40.36%) at minimum requiring time (18.27 days) including heavier sizes of callus (1.54 g) than BINA mung 7 when BINA mung 5 further recorded the longest root (2.92 cm) compare to BINA mung 7. Effect of treatments of the present study were significantly influenced the whole characters regarding callus culture, shoot regeneration and root proliferation. The highest percentage of callus (88.44%) within minimum time (12.53 days) including larger sizes callus (3.521 g) were produced in 1.0 mg L–1 BAP + 2.5 mg L–1 NAA among the treatments while the highest percentage of regenerated shoot (83.44%) at minimum requiring time (17.59 days) and more shoots (7.69 callus–1) were obtained in 1.0 mg L–1 BAP + 2.0 mg L– 1 NAA. Root induction (82.50%), number of roots plantlet–1 (8.469) with minimum requiring time for initiation (14.13 days) and root length (5.250 cm) were the highest in 0.2 mg L–1 IAA + 1.0 mg L–1 kinetin + 0.2 mg L–1 BAP. Incase of interaction, percentage of callus initiation (89.38 %) was the highest in BINA mung 5 treated by 1.0 mg L–1 BAP + 2.5 mg L–1 NAA at requiring minimum time (12.38 days) while same treatment produced the larger callus (3.581 g) among the interactions. The highest percentage (84.38%) and number (7.813 callus–1) of shoot with minimum requiring time (17.50 days) were found from BINA mung 5 treated by 1.0 mg L–1 BAP + 2.0 mg L–1 NAA. Similarly, the longest shoot (5.58 cm) was produced from the BINA mung 5 treated by 1.0 mg L–1 BAP + 2.0 mg L–1 NAA. However, root induction (%), roots plantlet–1, days required for root initiation and root length were statistically similar among the whole interaction treatments due to non significant variation. This result mentioned that the variety BINA mung 5 was better than BINA mung 7 for callus induction, shoot regeneration and root initiation while 1.0 mg L–1 BAP + 2.5 mg L–1 NAA, 1.0 mg L–1 BAP + 2.0 mg L–1 NAA and 0.2 mg L–1 IAA + 1.0 mg L–1 kinetin + 0.2 mg L–1 BAP supplemented with MS medium were the best combinations for better callusing, higher ability of shoot regeneration and root proliferation.DOI: http://dx.doi.org/10.3329/jesnr.v7i2.22203 J. Environ. Sci. & Natural Resources, 7(2): 47-52 2014

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.

scholarly journals Morphological and Histological Evaluations of In Vitro Regeneration in Elliottia racemosa Leaf Explants Induced on Media with Thidiazuron

2008 ◽  
Vol 133 (2) ◽  
pp. 167-172 ◽  
Author(s):  
Seong Min Woo ◽  
Hazel Y. Wetzstein
Keyword(s):  
Shoot Organogenesis ◽  
In Vitro Regeneration ◽  
Leaf Explants ◽  
Root Apex ◽  
Small Tree ◽  
Cell Layers ◽  
Scanning Electron Micrography ◽  
Indole 3 Acetic Acid ◽  
Deciduous Shrub

Georgia plume (Elliottia racemosa Muhlenb. ex. Elliott) is a rare deciduous shrub or small tree. It has sustained severe loss of habitat and its range is now restricted to a limited number of sites in the state of Georgia. Tissue culture protocols have been developed as a means to propagate and conserve this threatened species using leaf explants induced on medium supplemented with 10 μm thidiazuron (TDZ) and 5 μm indole-3-acetic acid (IAA). Bud-like clusters, elongated embryo-like protrusions, and shoot-like structures were produced from the leaf explants. Morphological and histological evaluations of cultures during induction and development were conducted using light microscopy of sectioned material and scanning electron micrography. Histology of explant tissues indicates that plant regeneration of Georgia plume occurs through a shoot organogenesis pathway that involves the formation of actively dividing meristematic regions originating in subepidermal cell layers that proliferate to form protuberances on the explant surface. Numerous well-formed shoot apical meristems with leaf primordia are produced, as well as fused shoot-like structures. Elongated, embryo-like structures had various degrees of shoot apex development. Evaluations of serial sections found that they lacked a defined root apex, and that basal portions were composed of parenchymatous files of cells with a broad point of attachment to the parent tissue. The lack of bipolarity and a root pole signifies that true somatic embryogenesis does not occur.

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