Advertisement

Springer Nature is making Coronavirus research free. View research | View latest news | Sign up for updates

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

  • 406 Accesses

  • 10 Citations

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.

This is a preview of subscription content, log in to check access.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

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

References

  1. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein dye binding. Anal Biochem 72:248–254. doi:10.1016/0003-2697(76)90527-3

  2. Chilton MD, Tepfer DA, Petit A, David C, Casse-Delbart F, Tempe J (1982) Agrobacterium rhizogenes inserts T-DNA into the genome of the host plant root cells. Nature 295:432–434. doi:10.1038/295432a0

  3. Cho H-J, Widholm JM, Tanaka N, Nakanishi Y, Murooka Y (1998) Agrobacterium rhizogenes-mediated transformation and regeneration of the legume Astragalus sinicus (Chinese milk vetch). Plant Sci 138:53–65. doi:10.1016/S0168-9452(98)00149-6

  4. Cho H-J, Farrand SK, Noel GR, Widholm JM (2000) High efficiency induction of soybean hairy roots and propagation of the soybean cyst nematode. Planta 210:195–204. doi:10.1007/PL00008126

  5. Chung KJ, Hwang SK, Hahn BS, Kim KH, Kim JB, Kim YH, Yang JS, Ha SH (2008) Authentic seed-specific activity of the Perilla oleosin 19 gene promoter in transgenic Arabidopsis. Plant Cell Rep 27:29–37. doi:10.1007/s00299-007-0440-6

  6. Costantino P, Spano L, Pomponi M, Benvenuto E, Ancora G (1984) The T-DNA of Agrobacterium rhizogenes is transmitted through meiosis to the progeny of hairy root plants. J Mol Appl Genet 2:465–470

  7. Guillon S, Tremouillaux-Guiller J, Pati PK, Rideau M, Gantet P (2006) Harnessing the potential of hairy roots: dawn of a new era. Trends Biotechnol 24:403–409. doi:10.1016/j.tibtech.2006.07.002

  8. Hernalsteens J-P, Van Vliet F, De Beuckeleer M, Depicker A, Engler G, Lemmers M, Holsters M, Van Montagu M, Schellet J (1980) The Agrobacterium tumefaciens Ti plasmid as a host vector system for introducing foreign DNA in plant cells. Nature 287:654–657. doi:10.1038/287654a0

  9. Karimi M, Inze D, Depicker A (2002) Gateway vectors for Agrobacterium-mediated plant transformation. Trends Plant Sci 7:193–195. doi:10.1016/S1360-1385(02)02251-3

  10. Kim DG, Ferris H (2002) Relationship between crop losses and initial population densities of Meloidogyne arenaria in winter-grown Oriental melon in Korea. J Nematol 34:43–49

  11. Matsuda Y, Toyoda H, Sawabe A, Maeda K, Shimizu N, Fujita N, Fujita T, Nonomura T, Ouchi S (2000) A hairy root culture of melon produces aroma compounds. J Agric Food Chem 48:1417–1420. doi:10.1021/jf9906580

  12. Mugnier J (1987) Infection by Polymyxa betae and Plasmodiophora brassicae of roots containing root inducing transferred DNA of Agrobacterium rhizogenes. Phytopathology 77:539–542. doi:10.1094/Phyto-77-539

  13. Mugnier J, Mosse B (1987) Vesiculararbuscular mycorrhizal infection in transformed root inducing T-DNA roots grown axenically. Phytopathology 77:1045–1050. doi:10.1094/Phyto-77-1045

  14. Mugnier J, Ready PW, Riedeh GE (1986) Root culture system useful in the study of biotrophic root pathogens in vitro. In: Agustine PC, Danforth HD, Bakst MR (eds) Biotechnology for solving agricultural problems. Martinus Nijhoff Publishers, Dordrecht, pp 147–153

  15. Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:473–497. doi:10.1111/j.1399-3054.1962.tb08052.x

  16. Schröder J, Schröder G, Huisman H, Schilperoort RA, Schell J (1981) The mRNA for lysopine dehydrogenase in plant. Tumor cells is complementary to a Ti-plasmid fragment. FEBS Lett 129:166–168. doi:10.1016/0014-5793(81)80781-8

  17. Sijmons PC, Grundler FMW, von Mende N, Burrows PR, Wyss U (1991) Arabidopsis thaliana as a new model host for plant-parasitic nematodes. Plant J 1:245–254. doi:10.1111/j.1365-313X.1991.00245.x

  18. Tepfer D (1984) Transformation of several species of higher plants by Agrobacterium rhizogenes: sexual transmission of the transformed genotype and phenotype. Cell 37:959–967. doi:10.1016/0092-8674(84)90430-6

  19. Verdejo S, Jaffee BA, Mankau R (1988) Reproduction of Meloidogyne javanica on plant roots genetically transformed by Agrobacterium rhizogenes. J Nematol 20:599–604

  20. Verdejo-Lucas S, Pinochet J (1992) Population densities of five migratory endoparasitic nematodes in carrot diskcultures. J Nematol 24:96–98

Download references

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.

Author information

Correspondence to Bum-Soo Hahn.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Pak, H., Sim, J., 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

Download citation

Keywords

  • Oriental melon
  • Hairy root
  • A. rhizogenes
  • Root-knot nematode
  • Egg mass