scholarly journals Development of a protocol for genetic transformation of Malus spp

Caryologia ◽  
2021 ◽  
Vol 74 (3) ◽  
pp. 9-19
Author(s):  
Federico Martinelli ◽  
Anna Perrone ◽  
Abhaya M. Dandekar
Keyword(s):  
Genetic Transformation ◽  
Shoot Regeneration ◽  
Malus X Domestica ◽  
Selective Media ◽  
In Vitro Multiplication ◽  
Gene Construct

A protocol to produce transgenic shoots of Malus X domestica cv Greensleaves was optimized using two gene constructs previously used to create parthenocarpic tomato, Ino-IaaM and DefH9-IaaM. The aim was to obtain sufficient nº of transgenic shoots for in vitro multiplication, transfer to soil, grafting and testing for parthenocarpy in the next years. We investigated the effects of two modifications of a previous published protocol: 1) co-transformation with an Agrobacterium containing “VIP” genes in the gene construct and 2) two different hormones or hormone combinations. More shoot regeneration was obtained with a combination of three hormones (BA:NAA:TDZ) during co-cultivation instead of IBA and no co-transformation was performed using the VIP gene. For the DefH9-IaaM transgene, 21.04% regeneration was achieved for this treatment instead of 8.95% achieved with “IBA treatment” and 4.42% with the Agrobacterium co-transformation treatment. More shoot regeneration occurred with the combination of three hormones (BA:NAA:TDZ) instead of with only IBA and no co-transformation was performed using VIP gene. Experiments using Ino-IaaM confirmed the results shown for the DefH9-IaaM transgene. The regenerated shoots were multiplied in selective media containing kanamycin and roots were obtained. 

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 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 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 Rooting in Km selective media as efficient in vitro selection method for sunflower genetic transformation

2006 ◽  
Vol 9 (3) ◽  
pp. 0-0 ◽  
Author(s):  
Laura Mabel Radonic ◽  
Julian Marcos Zimmermann ◽  
Diego Zavallo ◽  
Nilda Lopez ◽  
Marisa Lopez Bilbao
Keyword(s):  
Genetic Transformation ◽  
In Vitro Selection ◽  
Selection Method ◽  
Selective Media

scholarly journals In Vitro Shoot Regeneration of Oil Seed Crop Sesamum indicum L. from Seedling Cotyledon Explant to Lay Ground for Genetic Transformation in Ethiopia

2020 ◽  
Vol 2020 ◽  
pp. 1-10
Author(s):  
Micheale Yifter Weldemichael ◽  
Yemane Tsehaye Bayratsion ◽  
Desta Berhe Sbhatu ◽  
Girmay Gebresamuel Abraha ◽  
Hagos Mohammedseid Juhar ◽  
...  
Keyword(s):  
Genetic Transformation ◽  
Shoot Regeneration ◽  
Agricultural Research ◽  
Culture Conditions ◽  
Shoot Length ◽  
Leaf Number ◽  
Naphthaleneacetic Acid ◽  
Root Induction ◽  
Shoot Number

This study was conducted to develop an efficient regeneration protocol used for sesame genetic transformation. Published regeneration methods using 6-benzylaminopurine (BAP), indol-3-butyric acid (IBA), and α-naphthaleneacetic acid (NAA) were used in this experiment. Cotyledon explants of 14 Ethiopian genotypes collected from Humera Agricultural Research Center (HuARC) were used. Optimized culture conditions were used to investigate the regeneration response of different genotypes. Significant interactions between hormone treatments, MS media, and genotypes for shoot and root regeneration were recorded. Results showed that culture of cotyledons were developed and used as a source of explants for shoot regeneration. The highest shoot number, leaf number, and shoot length were recorded on explants cultured on 1.00 mg·L−1 BAP + 1.00 mg·L−1 NAA in Hirhir and Aberghele, 0.75 mg·L−1 BAP + 1.00 mg·L−1 NAA in Baha Zeyit, and 1.0 mg·L−1 BAP + 1.00 mg·L−1 NAA in Humera 1, respectively. The lowest shoot number, leaf number, and shoot length were observed on explants cultured on the control in Gondar 1, Borkana, and Baha Necho, Borkana and Beha Necho, and Baha Necho, respectively. Genotypes with well-developed shoots were transferred to root induction media. Under rooting media, the best mean, root number, and root length were observed in Aberghele and ACC44 at 0.1 mg·L−1 IBA and 0.2 mg·L−1 NAA, respectively. Standardizing genotypes trait data to mean 0 and unity variance has helped to group 14 genotypes into four distinct clusters. Hirhir, Abeghele, Baha Zeyit, and Humera 1 were the best genotypes. These findings lay ground to Ethiopian sesame to do further genetic transformation aiming at improving the crop.

scholarly journals IN VITRO SHOOT REGENERATION OF RUBUS PLANTS USING PETIOLES AND INTERNODES.

HortScience ◽  
1990 ◽  
Vol 25 (9) ◽  
pp. 1137d-1137
Author(s):  
Mahmoud A. Hassan ◽  
Harry Jan Swartz
Keyword(s):  
Genetic Transformation ◽  
Shoot Regeneration ◽  
Plantlet Regeneration ◽  
Ms Medium ◽  
In Vitro Shoot Regeneration

An efficient protocol for plantlet regeneration using petioles and internodes of two blackberry cultivars has been developed for use in genetic transformation. Maximum shoot regeneration was induced on MS medium supplemented with 5 μM Thidiazuron (TDZ) and 0.5 μM IBA. Preconditioning the source shoots on 0.5 μM TDZ containing MS medium did increase the number of regenerated shoots/explant, but did not affect the regeneration percentage. The antibiotic, kanamycin, significantly reduced the growth and regeneration of explants at 10 mg/L or higher. In contrast, cefatoxime at 100-500 mg/L increased explant growth and percentage regeneration.

scholarly journals IN VITRO CALLUS INITIATION AND DEVELOPMENT OF MEDICAGO

HortScience ◽  
1996 ◽  
Vol 31 (5) ◽  
pp. 755c-755
Author(s):  
Guochen Yang ◽  
Marihelen Kamp-Glass
Keyword(s):  
Genetic Transformation ◽  
Shoot Regeneration ◽  
Shoot Organogenesis ◽  
Callus Initiation ◽  
Callus Production ◽  
Callus Regeneration ◽  
Genetic Transformation System ◽  
Further Development ◽  
In Vitro Shoot Regeneration

The success of genetic transformation depends on the efficiency and rehability of in vitro shoot regeneration. This research was pursued to investigate how different plant growth regulators influence alfalfa callus initiation and development, thus to establish a foundation for further development of an efficient shoot organogenesis protocol for the genetic transformation system. BA, zeatin, and 2,4,5-trichlorophenoxyacetic acid (2,4,5-T) were evaluated for callus initiation and development. BA at 1 or 5 mg·liter–1, or zeatin at 5 mg·liter–1 promoted callus regeneration and further development toward shoot organogenesis. However, 2,4,5-T at 1 mg·liter–1 enhanced only callus production. These results can and will be used for further development of a shoot regeneration protocol to assist alfalfa genetic transformation.

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