scholarly journals Genetic transformation of new perspective winter wheat genotypes in vitro

2020 ◽  
Vol 26 ◽  
pp. 270-275
Author(s):  
L. V. Slivka ◽  
O. V. Dubrovna
Keyword(s):  
Transgenic Plants ◽  
Genetic Transformation ◽  
Winter Wheat ◽  
Target Gene ◽  
Callus Cultures ◽  
Genetic Construct ◽  
Wheat Genotypes ◽  
Agrobacterium Mediated Transformation ◽  
New Perspective

Aim. Optimization of conditions for genetic transformation of new perspective winter wheat genotypes and production of transgenic plants. Methods. Agrobacterium-mediated transformation in vitro culture using callus cultures. Results. The influence of the optical density of cells of agrobacterial suspension, the concentration of the antibiotic cefotaxime, the duration of coculture on the frequency of obtaining kanamycin-resistant regenerants of new winter wheat genotypes by genetic transformation of callus cultures were investigated. By Agrobacterium-mediated transformation of morphogenic calluses of new perspective winter wheat genotypes were obtained plant-regenerants in the genome of which revealed the complete incorporation of a genetic construct containing oat and nptII transgenes. Conclusions. Agrobacterium-mediated transformation of callus cultures of new perspective winter wheat genotypes was optimized, and transgenic plants with the target gene of ornithine-δ-aminotransferase were obtained. Keywords: Triticum aestivum, Agrobacterium-mediated transformation, callus cultures, ornithine-δ-aminotransferase gene.

scholarly journals Optimization of Agrobacterium-mediated transformation of perspective winter wheat genotypes in vitro

2020 ◽  
Vol 26 ◽  
pp. 190-195
Author(s):  
O. V. Dubrovna ◽  
L. V. Slivka
Keyword(s):  
Triticum Aestivum ◽  
Genetic Transformation ◽  
Winter Wheat ◽  
Optical Density ◽  
Callus Cultures ◽  
Optimal Parameters ◽  
Wheat Genotypes ◽  
Agrobacterium Mediated Transformation ◽  
New Perspective

Aim. Optimization of conditions for genetic transformation of new perspective winter wheat genotypes. Methods. Agrobacterium-mediated transformation in culture in vitro. Results. The influence of the optical density of cells of the agrobacterial suspension, the concentration of the antibiotic cefotaxime, the duration of coculture on the frequency of obtaining kanamycin-resistant regenerants of new winter wheat genotypes by genetic transformation of callus cultures were investigated using LBA4404 and AGL0 strains. It is shown that depending on the strain the most optimal is the concentration of agrobacteria 0.2-03 OD, duration of coculture for 2-3 days and the use of cefotaxime at a concentration of 250-500 mg/L. Conclusions. The optimal parameters for conducting Agrobacterium-mediated transformation of callus cultures of new perspective winter wheat genotypes were selected. Keywords: Triticum aestivum, Agrobacterium-mediated transformation, callus cultures.

scholarly journals Receiving of genetic-modified wheat plants with heterolous ornitin-δ-aminotransferase gene

2018 ◽  
Vol 22 ◽  
pp. 222-227
Author(s):  
O. M. Honcharuk ◽  
O. V. Dubrovna
Keyword(s):  
Triticum Aestivum ◽  
Transgenic Plants ◽  
Bread Wheat ◽  
Binary Vector ◽  
Triticum Aestivum L ◽  
Callus Cultures ◽  
Pcr Analysis ◽  
Genetic Construct ◽  
Agrobacterium Mediated Transformation

Aim. Receiving of genetically modified plants of bread wheat with heterologous ornithine‑δ‑aminotransferase gene. Methods. Agrobacterium-mediated transformation of callus cultures in vitro, PCR-analysis. Results. By Agrobacterium-mediated transformation of the morphogenic calluses of bread wheat (Triticum aestivum L.) using the AGLO strain containing the binary vector pBi-OAT with the target ornithine-δ-aminotransferase (oat) and selective neomycinphosphotransferase II (nptII), transgenic plants-regenerators have been obtained. Conclusions. As a result of the genetic transformation of Zimoyarka variety, 12 wheat regenerants were obtained in the genome which revealed a complete integration of the genetic construct containing the oat and nptII transgenes. Keywords: Triticum aestivum L., Agrobacterium-mediated transformation, ornithine‑δ‑aminotransferase gene, PCR-analysis.

scholarly journals Agrobacterium-mediated in planta genetic transformation of winter wheat (Triticum aestivum L.)

2018 ◽  
Vol 22 ◽  
pp. 293-298
Author(s):  
S. I. Mykhalska ◽  
A. G. Komisarenko ◽  
V. M. Kurchii ◽  
O. M. Tishchenko
Keyword(s):  
Triticum Aestivum ◽  
Transgenic Plants ◽  
Genetic Transformation ◽  
Winter Wheat ◽  
Natural Frequency ◽  
Triticum Aestivum L ◽  
Transformation Method ◽  
In Planta ◽  
Agrobacterium Mediated Transformation ◽  
Pollination Methods

Aim. To optimize the agrobacterium-mediated method of winter wheat transformation (Triticum aestivum L.); to select the conditions and period of inoculation to effectively transfer the genes during pollination. Methods. Agrobacterium-mediated in planta genetic transformation of winter wheat (Triticum aestivum L.) during pollination. Results. The conditions for agrobacterium-mediated transformation method of winter wheat during natural (frequency pollination was 1 %) and non-natural (frequency pollination was 4 %) pollination were defined. Conclusions. The possibility of integrating transgenes into the genome of winter wheat plants by the method of Agrobacterium-mediated transformation in planta in the process of forced and natural pollination is demonstrated. It is found that the transformation efficiency to a large extent depends on the plant genotype and the method of carrying out the transformation procedure. The selection of transgenic plants under water deficit conditions allowed to identify the plants with functional transgene. The signs of functioning transgene have been remaining in the next generation of genetically modified winter wheat. Keywords: Triticum aestivum L., Agrobacterium-mediated transformation in planta, transgenic plants, seeds.

scholarly journals Tissues of immature and mature embryos as morphogenic competent explants for genetic transformation of wheat

2020 ◽  
Vol 26 ◽  
pp. 233-238
Author(s):  
S. I. Mykhalska ◽  
A. G. Komisarenko ◽  
V. M. Kurchii
Keyword(s):  
Triticum Aestivum ◽  
Winter Wheat ◽  
Molecular Genetic ◽  
Triticum Aestivum L ◽  
Callus Cultures ◽  
Molecular Genetic Analysis ◽  
Pcr Analysis ◽  
Agrobacterium Mediated Transformation ◽  
Mature Embryos

Aim. To analyze the efficiency of using tissues of immature and mature embryos for Agrobacterium-mediated transformation in vitro of new breeding-valuable genotypes of winter wheat (Triticum aestivum L.) for the purpose of their genetic improvement. Methods. Culture of in vitro, extraction and electrophoresis of DNA, PCR analysis. Results. The efficiency of induction of callusogenesis and regeneration of winter wheat shoots was analyzed. The morphogenetic response of callus cultures obtained from different explants under Agrobacterium-mediated transformation in vitro was investigated. Molecular genetic analysis of wheat regenerants for transgenes was performe. Conclusions. The tissues of immature and mature embryos of novel breeding-valuable wheat genotypes are competent explants for Agrobacterium-mediated transformation in vitro. In this case, the tissues of two daily sprouts of mature wheat germ are characterized by higher morphogenetic parameters, which helps to obtain a greater percentage of genetically modified variants. Keywords: Triticum aestivum L., in vitro, Agrobacterium-mediated transformation, immature embryos, mature embryos.

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.

scholarly journals Glyphosate selection of maize transformants containing CP4epsps gene

2020 ◽  
Vol 26 ◽  
pp. 239-244
Author(s):  
I. O. Nitovska ◽  
B. V. Morgun ◽  
O. Ye. Abraimova ◽  
T. M. Satarova
Keyword(s):  
Transgenic Plants ◽  
Dna Analysis ◽  
Selective Agent ◽  
Regeneration Rate ◽  
Agrobacterium Mediated Transformation ◽  
Plant Dna ◽  
Morphogenic Callus ◽  
Selection Of ◽  
Maize Callus

Aim. To study the selection conditions of maize transformants containing the CP4epsps gene using glyphosate as a selective agent. Methods. Tissue culture in vitro, Agrobacterium-mediated transformation, selection of transgenic plants, isolation of total plant DNA, analysis of plant DNA by polymerase chain reaction (PCR). Results. The morphogenic maize callus of immature embryos of the hybrid (PLS61)R2×PLS61 was produced, which had a high regeneration rate (up to 95%), that persisted over long cultivation. Agrobacterium mediated transformation of the morphogenic callus and selection of the transgenic material using glyphosate yielded maize transformants containing the CP4epsps gene at a frequency of 1%. Conclusions. Maize genotype (PLS61)R2×PLS61 is promising for studies on the maize genetic transformation, in particular for the production of transgenic maize resistant to glyphosate herbicide. The use of morphogenic maize callus (PLS61)R2×PLS61 and glyphosate as a selective agent at a concentration of 0.1 mM and 0.25 mM in media for callusogenesis and 0.01 mM in the medium for regeneration was effective for the selection of transgenic plants with the gene CP4epsps. Keywords: Zea mays L., morphogenic callus, Agrobacterium-mediated transformation, PCR, genetic engineering.

scholarly journals Genetic transformation of promising genotypes of winter bread wheat by in planta method

2021 ◽  
Vol 28 ◽  
pp. 106-111
Author(s):  
L. V. Slivka ◽  
O. V. Dubrovna
Keyword(s):  
Triticum Aestivum ◽  
Transgenic Plants ◽  
Genetic Transformation ◽  
Bread Wheat ◽  
Optical Density ◽  
Triticum Aestivum L ◽  
In Planta ◽  
Agrobacterium Mediated Transformation ◽  
Winter Bread Wheat ◽  
Inoculation Medium

Aim. Optimization of conditions and genetic transformation of new promising genotypes of winter bread wheat (Triticum aestivum L.) by in planta method. Methods. Agrobacterium-mediated transformation by the in planta method using strain AGL0 and vector construct pBi-OAT. Results. The influence of air temperature, optical density of cells of agrobacterial suspension, inoculation day and composition of inoculation medium on the frequency of obtaining transgenic plants of new promising genotypes of winter wheat was studied. The dependence of the frequency of obtaining transgenic plants on environmental conditions, in particular temperature, has been established. It was found that the temperature regime of 20-22 °C provided the largest number (4.4%) of wheat transformants, and when the temperature is reduced to 16-18 °C there is a decrease in the efficiency of T-DNA transfer into the plant genome and the lowest frequency of transformation is observed. Conclusions. The largest number of transformants was obtained using a inoculation medium without sucrose, the optical density of cells of the agrobacterial suspension of 0.4 op.od. and inoculation on the third day after castration of ears. Keywords: Triticum aestivum L., Agrobacterium-mediated transformation, ornithine-δ-aminotransferase gene.

scholarly journals Standardized Genetic Transformation Protocol for Chrysanthemum cv. ‘Jinba’ with TERMINAL FLOWER 1 Homolog CmTFL1a

Genes ◽  
2020 ◽  
Vol 11 (8) ◽  
pp. 860
Author(s):  
Saba Haider ◽  
Yaohui Gao ◽  
Yike Gao
Keyword(s):  
Transgenic Plants ◽  
Genetic Transformation ◽  
Model Organism ◽  
Plant Morphology ◽  
Pcr Analysis ◽  
Transformation System ◽  
Transformation Protocol ◽  
Genetic Transformation System ◽  
Chrysanthemum X Morifolium

Chrysanthemum (Chrysanthemum x morifolium Ramat.) cultivar Jinba is a distinctive short-day chrysanthemum that can be exploited as a model organism for studying the molecular mechanism of flowering. The commercial value of Jinba can be increased in global flower markets by developing its proper regeneration and genetic transformation system. By addressing typical problems associated with Agrobacterium-mediated transformation in chrysanthemum, that is, low transformation efficiency and high cultivar specificity, we designed an efficient, stable transformation system. Here, we identify the features that significantly affect the genetic transformation of Jinba and standardize its transformation protocol by using CmTFL1a as a transgene. The appropriate concentrations of various antibiotics (kanamycin, meropenem and carbenicillin) and growth regulators (6-BA, 2,4-D and NAA) for the genetic transformation were determined to check their effects on in vitro plant regeneration from leaf segments of Jinba; thus, the transformation protocol was standardized through Agrobacterium tumefaciens (EHA105). In addition, the presence of the transgene and its stable expression in CmTFL1a transgenic plants were confirmed by polymerase chain reaction (PCR) analysis. The CmTFL1a transgene constitutively expressed in the transgenic plants was highly expressed in shoot apices as compared to stem and leaves. Overexpression of CmTFL1a led to a delay in transition to the reproductive phase and significantly affected plant morphology. This study will help to understand the biological phenomenon of TFL1 homolog in chrysanthemum. Moreover, our findings can explore innovative possibilities for genetic engineering and breeding of other chrysanthemum cultivars.

scholarly journals Agrobacterium-mediated Genetic Transformation of Lentil (Lens culinaris Medik.) with Chitinase Gene followed by In vitro Flower and Pod Formation

2019 ◽  
Vol 29 (1) ◽  
pp. 99-109
Author(s):  
Subroto K Das ◽  
Kishwar Jahan Shethi ◽  
MI Hoque ◽  
RH Sarker
Keyword(s):  
Genetic Transformation ◽  
Lens Culinaris ◽  
Target Gene ◽  
Gene Selection ◽  
Chitinase Gene ◽  
Pcr Analysis ◽  
Agrobacterium Strain ◽  
Half Strength ◽  
Selection Of

To investigate the integration of chitinase gene in lentil (Lens culinaris Medik.) namely, BARI masur-4 (BM-4), BARI masur-5 (BM-5) and BARI masur-6 (BM-6) through Agrobacterium-mediated genetic transformation was performed using Agrobacterium strain EHA 105 harboring bar (resistant to phosphinotrycin) and chitinase (gene of interest) gene. Selection of transformed shoots was carried out by gradually increasing the concentration of phosphinotrycin (PPT) up to 2.0 mg/l. Transgenic lentil shoots were produced with an overall frequency of 0.36 in case of BM-4 and BM-6 and 0.34 in case of BM-5, respectively. Most of the selected shoots developed in vitro flowers and pods following their sub-culture on half strength of MS supplemented with 20 mg/l IBA, 0.5 mg/l NAA with 50 mg/l ticarcillin. Seedlings germinated from the seeds were successfully transferred to soil for the development of further progeny. Stable integration of target gene was confirmed through PCR analysis. Plant Tissue Cult. & Biotech. 29(1): 99-109, 2019 (June)

scholarly journals Infection of Embryonic Callus with Agrobacterium Enables High-Speed Transformation of Maize

2019 ◽  
Vol 20 (2) ◽  
pp. 279 ◽  
Author(s):  
Dengxiang Du ◽  
Ruchang Jin ◽  
Jinjie Guo ◽  
Fangdong Zhang
Keyword(s):  
Transgenic Plants ◽  
Embryogenic Callus ◽  
High Speed ◽  
Molecular Breeding ◽  
Transformation Efficiency ◽  
Lytic Enzyme ◽  
Phenotypic Analysis ◽  
Agrobacterium Mediated Transformation ◽  
Prolonged Culture

Several approaches have recently been adopted to improve Agrobacterium-mediated transformation of maize; however, about eight months of in vitro culture are still required to isolate transgenic plants. Furthermore, genetic transformation of maize depends on immature embryos, which greatly increases costs. Here, we report a method that ensures the competency of an embryogenic callus secondary culture under laboratory conditions for Agrobacterium-mediated transformation. Moreover, pretreatment of the cell wall with a mixed lytic enzyme solution prior to Agrobacterium infection, significantly improved transformation efficiency and stability. Average stable transformation efficiency was approximately 30.39%, with peaks of 94.46%. Expression and phenotypic analysis of the Rsc reporter gene were tested in the T0 generation of transgenic plants. Using this system, we successfully regenerated transgenic maize plantlets within three months of the emergence of the embryogenic callus. Additionally, we reduced somaclonal variation accompanying prolonged culture of maize cells in the dedifferentiated state, thus facilitating the molecular breeding of maize.

Sign in / Sign up

Export Citation Format

Share Document