scholarly journals In vitro plant regeneration and Agrobacterium-mediated genetic transformation of a carnivorous plant, Nepenthes mirabilis

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Sissi Miguel ◽  
Cindy Michel ◽  
Flore Biteau ◽  
Alain Hehn ◽  
Frédéric Bourgaud
Keyword(s):  
Genetic Transformation ◽  
Shoot Regeneration ◽  
Carnivorous Plant ◽  
Feeding Strategies ◽  
In Vitro Plant Regeneration ◽  
Rt Pcr ◽  
Transformation Protocol ◽  
Positive Method ◽  
Stem Segments

Abstract In nutrient-poor habitats, carnivorous plants have developed novel feeding strategies based on the capture and digestion of prey and the assimilation of prey-derived nutrients by specialized traps. The Nepenthes genus, comprising nearly 160 species, presents a remarkable pitcher-shaped trap, leading to great interest among biologists, but the species of this genus are listed as threatened. In this work, we developed a protocol for reproducing Nepenthes mirabilis through shoot regeneration from calli. The cultivation of stem segments of N. mirabilis on MS medium containing thidiazuron induced organogenic calli after 10 weeks. Subcultured calli exposed to 6-benzylaminopurine showed shoot regeneration in 3 weeks with considerable yields (143 shoots/g of calli). Excised shoots transferred to medium with indole-3-butyric acid allowed rooting in 4 weeks, and rooted plantlets had a 100% survival rate. Based on this method, we also developed an Agrobacterium-mediated genetic transformation protocol using calli as explants and ipt as a positive method of selection. Twelve weeks post infection, regenerated shoots were observed at the surface of calli. Their transgenic status was confirmed by PCR and RT-PCR. In conclusion, this study provides an efficient method for regenerating Nepenthes and the first protocol for its stable genetic transformation, a new tool for studying carnivory.

scholarly journals Agrobacterium tumefaciens mediated transformation of the aquatic carnivorous plant Utricularia gibba

2020 ◽  
Author(s):  
Araceli Oropeza-Aburto ◽  
Sergio Alan Cervantes-Perez ◽  
Victor A Albert ◽  
Luis Rafael Herrera-Estrella
Keyword(s):  
Agrobacterium Tumefaciens ◽  
Genetic Transformation ◽  
In Vitro Selection ◽  
Carnivorous Plant ◽  
Carnivorous Plants ◽  
35S Promoter ◽  
Transformation Protocol ◽  
Utricularia Gibba

Abstract Background The genus Utricularia belongs to Lentibulariaceae, the largest family of carnivorous plants, which includes terrestrial, epiphytic and aquatic species. The development of specialized structures that evolved for carnivory is a feature of this genus that has been of great interest to biologists since Darwin‘s early studies. Utricularia gibba is itself an aquatic plant with sophisticated bladder traps having one of the most complex suction mechanisms for trapping prey. However, the molecular characterization of the mechanisms that regulate trap development and the biophysical processes involved in prey trapping are still largely unknown due to the lack of a simple and reproducible gene transfer system. Results Here, we report the establishment of a simple, fast and reproducible protocol for genetic transformation of U. gibba based on the T-DNA of Agrobacterium tumefaciens . An in vitro selection system using Phosphinotricin as a selective agent was established for U. gibba . Plant transformation was confirmed by histochemical GUS assays and PCR and qRT-PCR analyses. We report on the expression pattern of the 35S promoter and of the promoter of a trap-specific ribonuclease gene in transgenic U. gibba plants. Conclusions The genetic transformation protocol reported here is an effective method for studying developmental biology and functional genomics of this genus of carnivorous plants and advances the utility of U. gibba as a model system to study developmental processes involved in trap formation.

scholarly journals Agrobacterium tumefaciens mediated transformation of the aquatic carnivorous plant Utricularia gibba

2020 ◽  
Author(s):  
Araceli Oropeza-Aburto ◽  
Sergio Alan Cervantes-Perez ◽  
Victor A Albert ◽  
Luis Rafael Herrera-Estrella
Keyword(s):  
Agrobacterium Tumefaciens ◽  
Genetic Transformation ◽  
In Vitro Selection ◽  
Carnivorous Plant ◽  
Carnivorous Plants ◽  
35S Promoter ◽  
Transformation Protocol ◽  
Utricularia Gibba

Abstract Background The genus Utricularia belongs to Lentibulariaceae, the largest family of carnivorous plants, which includes terrestrial, epiphytic and aquatic species. The development of specialized structures that evolved for carnivory is a feature of this genus that has been of great interest to biologists since Darwin‘s early studies. Utricularia gibba is itself an aquatic plant with sophisticated bladder traps having one of the most complex suction mechanisms for trapping prey. However, the molecular characterization of the mechanisms that regulate trap development and the biophysical processes involved in prey trapping are still largely unknown due to the lack of a simple and reproducible gene transfer system. Results Here, we report the establishment of a simple, fast and reproducible protocol for genetic transformation of U. gibba based on the T-DNA of Agrobacterium tumefaciens . An in vitro selection system using Phosphinotricin as a selective agent was established for U. gibba . Plant transformation was confirmed by histochemical GUS assays and PCR and qRT-PCR analyses. We report on the expression pattern of the 35S promoter and of the promoter of a trap-specific ribonuclease gene in transgenic U. gibba plants. Conclusions The genetic transformation protocol reported here is an effective method for studying developmental biology and functional genomics of this genus of carnivorous plants and advances the utility of U. gibba as a model system to study developmental processes involved in trap formation.

scholarly journals Agrobacterium tumefaciens mediated transformation of the aquatic carnivorous plant Utricularia gibba

2020 ◽  
Author(s):  
Araceli Oropeza-Aburto ◽  
Sergio Alan Cervantes-Perez ◽  
Luis Rafael Herrera-Estrella
Keyword(s):  
Agrobacterium Tumefaciens ◽  
Genetic Transformation ◽  
In Vitro Selection ◽  
Carnivorous Plant ◽  
35S Promoter ◽  
Specific Expression ◽  
Transformation Protocol ◽  
Utricularia Gibba

Abstract Background Utricularia genus belongs to Lentibulariaceae family and has the highest number of species including terrestrials, epiphytes and aquatics plants. The development of specialized structures that evolved for carnivory is a feature of this genus that has been of great interest to biologist since the early studies of Darwin. Utricularia gibba is an aquatic carnivorous plant with sophisticated bladders traps that have one of the most complex suction mechanisms for trapping a prey. However, the molecular characterization of trap developmental and the biophysical processes involved in prey trapping are still largely unknown due to the lack of simple and reproducible gene transfer system for carnivorous plants.Results Here, we report the establishment of a simple, fast and effective protocol for the genetic transformation of U. gibba based on the T-DNA of Agrobacterium tumefaciens . An in vitro selection system using Phosphinotricin as selective agent was established for U. gibba. We report the tissue specific expression of the 35S promoter and the promoter of a trap specific ribonuclease gene. Plant transformation was confirmed by PCR and Real Time PCR in U. gibba plants.Conclusions We conclude that the genetic transformation protocol we developed is an effective method to study developmental biology and functional genomics of carnivory and propose U. gibba as model to study the developmental processes involved in trap formation.

scholarly journals Callus induction and regeneration of Alkanna orientalis var. Orientalis and A. sieheana

2019 ◽  
Vol 48 (3) ◽  
pp. 633-640 ◽  
Author(s):  
Cenney Yaman ◽  
Serkan Uranbey ◽  
Hussein Abdullah Ahmed ◽  
Sabahattin Özcan ◽  
Osman Tugay ◽  
...  
Keyword(s):  
Shoot Regeneration ◽  
Callus Induction ◽  
High Frequency ◽  
Stem Segment ◽  
Leaf Base ◽  
Cotyledonary Leaf ◽  
Frequency Variations ◽  
Alkanna Orientalis ◽  
Stem Segments

Callus induction and proliferation of Alkanna orientalis var. orientalis and Alkanna sieheana containing valuable alkannin/shikonin (A/S) derivates were investigated using leaf base and stem segment explants. Stem segments and cotyledonary leaf base of both species were cultured on Murashige and Skoog medium fortified with different concentrations of BAP, Kn, NAA, IAA and IBA for callus induction and shoot regeneration. High frequency reproducible, prolific and compact calli formation was obtained from the stem segments of both species in all media tested. The frequency variations of callus induction and shoot regeneration were discussed in terms of different species, plant growth regulators and explant resources. A. orientalis and A. sieheana may be considered to be alternative plants for the A/S production in vitro.

scholarly journals Direct Shoot Organogenesis of Medicago sativa

HortScience ◽  
1996 ◽  
Vol 31 (4) ◽  
pp. 628f-628
Author(s):  
Guochen Yang ◽  
Marihelen Kamp-Glass
Keyword(s):  
Genetic Transformation ◽  
Shoot Regeneration ◽  
Shoot Organogenesis ◽  
Multiple Shoots ◽  
Direct Shoot Regeneration ◽  
Callus Initiation ◽  
Direct Shoot Organogenesis ◽  
Direct Shoot ◽  
In Vitro Shoot Regeneration

An efficient and reliable protocol of in vitro shoot regeneration must be first established to have a successful genetic transformation. As a member of legume family, alfalfa is very difficult for direct shoot regeneration. There is no published information on direct shoot organogenesis, although success has been well documented on embryogenesis, which must go through callus stage. Different plant growth regulators at various concentrations were evaluated for callus initiation, development, and direct shoot regeneration. Multiple shoots were produced directly from each individual explant. This will provide an efficient means for production of transgenic alfalfa plants. Therefore, genetic transformation of Medicago germplasm will be significantly expedited.

scholarly journals Propagation of Sedum spectabile Boreau in Leaf Culture in Vitro

2012 ◽  
Vol 40 (1) ◽  
pp. 107 ◽  
Author(s):  
Cuiqin YANG ◽  
Yaoguo QIN ◽  
Xin SUN ◽  
Shu YUAN ◽  
Honghui LIN
Keyword(s):  
Shoot Regeneration ◽  
Culture Medium ◽  
Shoot Proliferation ◽  
Shoot Formation ◽  
Root Induction ◽  
Ms Medium ◽  
Shoot Induction ◽  
Leaf Base ◽  
Stem Segments

An efficient protocol was established for Sedum spectabile Boreau propagation. Various leaf parts were used as explants to regenerate plantlets, the stem segments of which were cultured for shoot proliferation and plantlet multiplication. The results showed that the leaf base was the optimal explant, as compared to both the middle and the top of leaves, for shoot formation. The highest shoot induction of 88.9% was observed on MS medium supplemented with 0.6 mg/l TDZ and 0.1 mg/l NAA. Hyperhydric leaves obtained in primary culture developed first into abnormal somatic embryos 10 days after subculture, and then into hyperhydric plantlets after an additional 10 days. The hyperhydric plantlets reversed to normal plantlets when plant growth regulators were removed from culture medium. Further, stem segments from reversed plantlets were used for shoot regeneration and root induction. Optimal shoot regeneration was obtained in MS medium containing 0.6 mg/l TDZ with 0.1 mg/l NAA. Root induction and root mean number were all higher on auxin-free medium than on medium containing auxins.

scholarly journals In vitro organogenesis and genetic transformation of mandarin cultivars

2019 ◽  
Vol 41 (2) ◽  
Author(s):  
Leonardo Soriano ◽  
Eveline Carla da Rocha Tavano ◽  
Marcelo Favaretto Correa ◽  
Ricardo Harakava ◽  
Beatriz Madalena Januzzi Mendes ◽  
...  
Keyword(s):  
Transgenic Plants ◽  
Genetic Transformation ◽  
Shoot Regeneration ◽  
Citrus Reticulata ◽  
Explant Type ◽  
In Vitro Organogenesis ◽  
Citrus Clementina ◽  
Number Of Shoots

Abstract The in vitro organogenesis of Fremont (Citrus clementina x ), Citrus reticulataThomas (Citrus reticulata), and Nules (Citrus clementina) mandarins was evaluated aiming to optimize a regeneration protocol that could be applied in genetic transformation. The use of epicotyl-derived explants resulted in higher explant responsiveness and number of shoots developed per explant when compared with the use of internodal-derived explants. The highest efficiency in shoot regeneration was observed in the presence of 1 mg L-1 of BAP, regardless of the explant type and cultivar. The in vitro organogenesis protocol produced transgenic plants from three mandarin cultivars expressing attA gene under the control of phloem-specific promoters.

Sign in / Sign up

Export Citation Format

Share Document