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.

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 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 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.

scholarly journals In Vitro Shoot Regeneration and Polyploid Induction from Leaves of Hypericum Species

HortScience ◽  
2009 ◽  
Vol 44 (7) ◽  
pp. 1957-1961 ◽  
Author(s):  
Elisabeth M. Meyer ◽  
Darren H. Touchell ◽  
Thomas G. Ranney
Keyword(s):  
Shoot Regeneration ◽  
Chromosome Doubling ◽  
In Vitro Regeneration ◽  
Leaf Explants ◽  
Shoot Induction ◽  
Bluish Green ◽  
Spontaneous Chromosome ◽  
Ornamental Traits ◽  
In Vitro Shoot Regeneration

Hypericum L. H2003-004-016 is a complex hybrid among Hypericum frondosum Michx., Hypericum galioides Lam., and Hypericum kalmianum L. and exhibits valuable ornamental characteristics, including compact habit, bluish green foliage, and showy flowers. Inducing polyploidy may further enhance the ornamental traits of this hybrid and provide new opportunities for hybridizing with other naturally occurring polyploid Hypericum sp. In this study, in vitro shoot regeneration and treatment of regenerative callus with the dinitroaniline herbicide oryzalin (3,5-dinitro-N4,N4-dipropylsufanilamide) were investigated as a means of inducing allopolyploidy. First, in vitro regeneration was optimized for callus and shoot induction by culture of leaf explants on medium supplemented with benzylamino purine (BA) or meta-topolin (mT) at 5, 10, or 15 μM in combination with indoleacetic acid (IAA) at 0, 1.25, 2.5, or 5 μM. Both BA and mT treatments successfully induced regenerative callus and shoots. Multiple regression analysis estimated maximum regenerative callus (94%) and shoot induction (18 shoots per explant) in medium supplemented with 5 μM BA and 3.75 μM IAA. In the second part of the study, exposure of regenerative callus to oryzalin at 0, 7.5, 15, 30, 60, or 90 μM for durations of 3, 6, or 9 d was investigated for polyploid induction. There was no survival for any of the calli in the 60- or 90-μM oryzalin treatments, but calli subjected to the other treatments exhibited some survival and polyploid induction. Duration had no effect on callus survival or ploidy level, but oryzalin concentration was a significant factor in both. The greatest percentage (44%) of polyploids was induced with 30 μM oryzalin. Spontaneous chromosome doubling was observed in 8% of control explants receiving no oryzalin treatment.

scholarly journals METHODS FOR PRODUCING GRAFT CHIMERAS IN VITRO

HortScience ◽  
1991 ◽  
Vol 26 (6) ◽  
pp. 756G-757 ◽  
Author(s):  
Huey-Jen Chen ◽  
Harry Jan Swartz
Keyword(s):  
Shoot Regeneration ◽  
Shoot Organogenesis ◽  
Woody Species ◽  
In Vitro Techniques ◽  
Golden Delicious ◽  
Periclinal Chimeras ◽  
Organogenesis In Vitro ◽  
Solanaceous Plants

Several authors report the synthesis of periclinal chimeras generated from graft unions of Solanaceous plants grown in the greenhouse. As this technique requires shoot organogenesis, in vitro conditions are necessary to adapt this technique to woody species. We now report several in vitro techniques necessary to mimic the in vivo graft chimera process. These include rootstock/scion preparation, micrografting and shoot organogenesis from graft unions. Zeatin and auxins have been helpful in preparing graftable material and for increasing the percentage successful grafts. A shorter exposure to organogenic medium containing thidiazuron resulted in greater percentage shoot regeneration from graft unions. Thorny/thornless Rubus and 'Liberty'/'Golden Delicious' or 'Gala' Malus (color) markers are being used to determine the percentage of these regenerants which are chimeral.

scholarly journals (405) Regeneration of Venus Fly Trap (Dionaea muscipula Ellis) from Leaf Culture

HortScience ◽  
2005 ◽  
Vol 40 (4) ◽  
pp. 1062F-1063
Author(s):  
Khalid M. Ahmad ◽  
Syed M. A. Zobayed ◽  
Praveen K. Saxena ◽  
David M. Hunter
Keyword(s):  
Somatic Embryogenesis ◽  
Shoot Organogenesis ◽  
In Vitro Regeneration ◽  
Leaf Disc ◽  
Leaf Explants ◽  
Carnivorous Plant ◽  
In Vitro Techniques ◽  
Direct Shoot Organogenesis ◽  
Direct Shoot

Dionaeamuscipula Ellis commonly known as Venus fly trap is an important carnivorous plant with medicinal importance. It contains certain secondary metabolites like naphthoquinones and is used in anti-aid and anti-cancer drugs and other medicines like Cornivora. Increasing interest and use as an ornamental and medicinal plant, and dietary supplement have put it in an endangered state. Development of in vitro techniques for the preservation of germplasm that is on the brink of extinction is highly demanded. A regeneration protocol for the multiplication and micropropagation of Dionaeamuscipla Ellis was established. In vitro regeneration potential of leaf explants in different concentrations and combinations of plant growth substances was investigated in this study. Seeds were grown and leaf disc explants were excised and cultured under aseptic conditions on nutritional medium containing half strength Murashige and Skoog (MS) mix with combinations of 1.0–20.0 μm BA, 2.5.0 μm IBA, 1.0–10.0 μm 2iP and 0.1–0.5μm TDZ. The cultures were kept in growth cabinet with cool white light (40–60 μmol·m-2·s-1) under 16-h photoperiod. Regeneration was recorded after 60 days with the intervals of 15 days based on the degree of shoot organogenesis and somatic embryogenesis. 1/2 MS + 0.1 TDZ appeared to be efficient for somatic embryogenesis and simple MS for direct shoot organogenesis. 1/2 MS combined with 2iP appeared to be efficient for regeneration either by direct shoot organogenesis or by somatic embryogenesis. Plants were rooted well in Cape Cundew medium. These investigations will aid in the development of a model system for clonal mass propagation and in vitro regeneration of Dionaeamuscipla Ellis.

scholarly journals Comparative Organogenic Response of Six Clonal Apple Rootstock Cultivars

HortScience ◽  
2016 ◽  
Vol 51 (3) ◽  
pp. 272-278 ◽  
Author(s):  
Qingrong Sun ◽  
Meijuan Sun ◽  
Hongyan Sun ◽  
Richard L. Bell ◽  
Linguang Li ◽  
...  
Keyword(s):  
Shoot Regeneration ◽  
Shoot Organogenesis ◽  
Leaf Explants ◽  
Basal Medium ◽  
Medium Composition ◽  
Apple Rootstock ◽  
Malus Prunifolia ◽  
Root Organogenesis ◽  
High Regeneration

The organogenesis potential is different among cultivars and must be optimized for individual genotype. Shoot organogenesis capacity from in vitro leaves and root organogenesis capacity of in vitro shoots in six clonal apple rootstock cultivars were compared. The shoot organogenesis capacity was highly genotype dependent. ‘GM256’ was found to be the most responsive genotype for shoot regeneration from leaf explants among the cultivars, showing high regeneration percentage on all tested media. The effects of basal medium composition and cytokinins on shoot regeneration were different depending on rootstock genotype. Optimum regeneration occurred on Murashige and Skoog (MS) basal medium for ‘71-3-150’, and optimum regeneration occurred on Quoirin and Lepoivre (QL) basal medium for ‘60-160’ and ‘ПБ’. Thidiazuron (TDZ) was more effective than 6-benzylaminopurine (BA) for Malus prunifolia (Y), whereas TDZ and BA were not significantly different for the other cultivars. All rootstock cultivars showed high root organogenic capacity. The percentage of rooting reached more than 90% and the mean root number per plantlet ranged from three to five. The optimum rooting medium was different for different rootstock cultivars. Optimum root organogenesis occurred on half-strength QL medium for ‘GM256’ and ‘Y’, and for ‘ПБ’ and ‘JM7’ on one-quarter-strength MS medium.

In vitro differentiation and plant regeneration from root and other explants of juvenile origin in pea (Pisum sativum L.)

2017 ◽  
Author(s):  
Shikha Sharma ◽  
Geetika Gambhir ◽  
D. K. Srivastava
Keyword(s):  
Pisum Sativum ◽  
Shoot Regeneration ◽  
In Vitro Regeneration ◽  
Leaf Explants ◽  
Cotyledonary Node ◽  
Shoot Elongation ◽  
Root Regeneration ◽  
Pisum Sativum L ◽  
Regenerated Plantlets

In vitro regeneration of pea explants (Pisum sativum L. var. ‘Lincon’) was done in 49 different combinations and concentrations of BAP, BAP and NAA, BAP and IBA, TDZ, TDZ and Adenine for shoot regeneration from hypocotyl, root, leaf and cotyledonary node. High frequency shoot regeneration was obtained in hypocotyl (81.43%), root(83.53%) and cotyledonary node(72.76%) on MS medium supplemented with 4.50 mg/l BAP and 1.86mg/l NAA, 2.00mg/l TDZ and4.50 mg/l BAP and 1.86mg/l NAA respectively. No shoot regeneration was obtained from leaf explants on any of the combination used. Shoot elongation was observed on the same medium used for shoot regeneration respectively.MS medium supplemented with 0.20 mg/l IBA was found best for root regeneration from in vitro raised shoots. The plantlets were able to regenerate within 6-7 weeks. The regenerated plantlets were acclimatized in pre-sterilized cocopeat.

scholarly journals Factors Influencing Shoot Regeneration and ß-Glucuronidase Expression from `Royal Gala' Apple Internodes

HortScience ◽  
1997 ◽  
Vol 32 (3) ◽  
pp. 442B-442
Author(s):  
Qingzhong Liu ◽  
Sarbagh Salih ◽  
Freddi Hammerschlag
Keyword(s):  
Disease Resistance ◽  
Agrobacterium Tumefaciens ◽  
Shoot Regeneration ◽  
Leaf Explants ◽  
Transformation Process ◽  
Factors Influencing ◽  
In Vitro Shoots ◽  
Light Green ◽  
Number Of Shoots

Factors influencing regeneration and ß-glucuronidase expression from apple (Malus × domestica Borkh.) stem internodes were studied as part of a program to develop transgenic `Royal Gala' apple with improved disease resistance. The early stages of the transformation process were monitored by counting the number of ß-glucuronidase (GUS) expressing zones immediately after co-cultivation of explants with Agrobacterium tumefaciens supervirulent strain EHA105 (p35SGUS_INT) and by counting the number of GUS-expressing calli developing on explants 2 weeks after co-cultivation. Etiolated shoots were produced from in vitro shoots cultured for 2 weeks in the light followed by 2 weeks in the dark and were compared with shoots cultured for 4 weeks in the light (green shoots). First internodes from etiolated shoots produced three, 10 and 100 times the number of shoots regenerated from second, third, and fourth internodal explants, respectively, and produced seven times the number of shoots compared with similar explants from green shoots. 100% of first internodes from etiolated shoots exhibited GUS-expressing zones and yielded twice as many GUS-expressing zones when compared with leaf explants from green shoots, which exhibited GUS-expressing zones in only 60% of the explants. An average of nine GUS-expressing calli per explant were produced on first internodes from etiolated shoots 2 weeks after co-cultivation.

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