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Elimination of macro elements from inoculation and co-cultivation media enhances the efficiency of Agrobacterium-mediated transformation in Petunia

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Abstract

In order to evaluate the effect of inoculation and co-cultivation media elements on transformation frequency in Petunia hybrida, modified MS media with different elements were tested on Alvan and Large Flower Alvan (LF Alvan), two local cultivars. Leaf explants of both cultivars were inoculated with Agrobacterium tumefaciens strain LBA4404 (pBI121) containing neomycin phosphotransferase (nptII) and an intron-containing β-glucuronidase (gus) genes. When medium lacking KH2PO4, NH4NO3, KNO3, and CaCl2 was used as inoculation and co-cultivation medium, a higher frequency of transformation for Alvan (22%) and LF Alvan (16%) was obtained. Kanamycin resistant plantlets were stained blue by GUS assay. Furthermore, polymerase chain reaction (PCR) analysis revealed the presence of both gus and nptII genes in all putative transformants. Finally, southern blot hybridization confirmed insertion of 1–4 copies of gus gene in transgenic plants.

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Abbreviations

LF Alvan:

Large Flower Alvan

GUS:

β-glucuronidase

CTAB:

Cetyltrimethylammonium bromide

nptII :

Neomycin phosphotransferase

CaMV:

Cauliflower mosaic virus

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Acknowledegments

This work was supported by Bu-ali Sina University, Hamedan, Iran, and Novin Giti Gene Biotech. Co. Biotechnology Incubator Center of National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran.

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Authors and Affiliations

  1. Department of Biotechnology, Faculty of Agriculture, Bu Ali-Sina University, Hamedan, 6517838695, Iran

    Aso Nobakht Vakili

  2. Department of Plant Biotechnology, Faculty of Agriculture, Bu Ali-Sina University, Hamedan, 6517838695, Iran

    Hedayat Bagheri

  3. Department of Genetic Engineering, Agricultural Biotechnology Research Institute of Iran (ABRII), Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran

    Pejman Azadi

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  1. Aso Nobakht Vakili
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  2. Hedayat Bagheri
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Correspondence to Hedayat Bagheri.

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Nobakht Vakili, A., Bagheri, H. & Azadi, P. Elimination of macro elements from inoculation and co-cultivation media enhances the efficiency of Agrobacterium-mediated transformation in Petunia. Physiol Mol Biol Plants 24, 703–710 (2018). https://doi.org/10.1007/s12298-018-0553-0

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