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Hairy root induction in Oriental melon (Cucumis melo) by Agrobacterium rhizogenes and reproduction of the root-knot nematode (Meloidogyne incognita)

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Abstract

Cotyledon explants of two Oriental melons produced hairy roots when cultured on Murashige and Skoog basal medium after infection by the Agrobacterium rhizogenes. Hairy roots were produced from the wounded surface of the cotyledon explants of Cucumis melo L. cv. Geumssaragi-euncheon on Murashige and Skoog selective medium and 86% of the GUS stained hairy roots were positive for the expression of beta-glucuronidase. The insertion of the gfp-gus fusion gene in the genomic DNA and the presence of the gfp-gus-specific transcript in the total RNAs of transgenic hairy roots were confirmed by PCR and RT-PCR, respectively. An immunoblot analysis of the transgenic hairy root extract revealed 97 kDa single bands coincident with the molecular weight of the GFP-GUS fusion proteins. ELISA demonstrated that the highest level of GFP-GUS fusion protein expression was 0.47% of the total soluble protein in a transgenic hairy root. The MS medium showed the fastest growth among three media types tested. Infection of the hairy roots with a root-knot nematode resulted in the development of a mature egg mass about 4–5 weeks after inoculation. The highest number of egg mass was obtained on the hairy roots cultured in SH medium containing 0.3% agar.

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Abbreviations

B5:

Gamborg B5

CAPS:

N-Cyclohexyl-3-aminopropanesulfonic acid

ELISA:

Enzyme-linked immunosorbent assay

GFP:

Green fluorescent protein

GUS:

Beta-glucuronidase

M-MLV RT:

Moloney murine leukemia virus reverse transcriptase

MS:

Murashige and Skoog

SH:

Schenk & Hildebrandt

TSP:

The total soluble protein

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Acknowledgments

We are grateful to Drs. Dong-Ro Choi and Hye–Rim Hahn (National Academy of Agricultural Science) for providing M. incognita. We thank Ms. Mi-Ra Yang for technical assistance. This work was supported by the grants (code no. 20080401034031) from BioGreen21 program and National Academy of Agricultural Science, Rural Development Administration, Korea.

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Authors and Affiliations

  1. National Academy of Agricultural Science, Suwon, 441-857, Korea

    Hyo-Kyung Pak, Joon-Soo Sim, Hyo-Rim Ko, Sun-Hwa Ha, Mun-Sup Yoon, Yong-Hwan Kim & Bum-Soo Hahn

  2. Department of Genetic Engineering, Sungkyunkwan University, Suwon, 440-746, Korea

    Yong Rhee & Sukchan Lee

  3. Jeollabuk-Do Agricultural Research and Extension Services, Iksan, 570-704, Korea

    Chan-Ho Kang

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  1. Hyo-Kyung Pak
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Correspondence to Bum-Soo Hahn.

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Pak, HK., Sim, JS., Rhee, Y. et al. Hairy root induction in Oriental melon (Cucumis melo) by Agrobacterium rhizogenes and reproduction of the root-knot nematode (Meloidogyne incognita). Plant Cell Tiss Organ Cult 98, 219–228 (2009). https://doi.org/10.1007/s11240-009-9556-4

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  • DOI: https://doi.org/10.1007/s11240-009-9556-4

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