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  • Citation:

    Cheng, Y.H., Yang, T.A., Yeh, S.D. (1996). Efficient transformation of papaya by coat protein gene of Papaya ringspot virus mediated by Agrobacterium following liquid-phase wounding of embryogenic tissues with caborundum. Plant Cell Reports, 16, 127-132.

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 peer-reviewed article

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 180308 LH: Standardized citation to APA style.

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Publication Year:

 1996

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  • Abstract:

    Generation of transgenic papaya (_Carica papaya_ L.) has been hampered by the low rates of transformation achieved by conventional Agrobacterium infection or microprojectile bombardment. We describe an efficient Agrobacterium-mediated transformation method based on wounding of cultured embryogenic tissues with carborundum in liquid phase. Embryogenic tissues were obtained from cultured immature zygotic embryos collected 75-90 days after pollination. The expressible coat protein (CP) gene of a Taiwan strain of Papaya ringspot virus (PRSV) was constructed in a Ti binary vector pBGCP, which contained the nptII gene as a selection marker. The embryogenic tissues were vortexed with 600 mesh carborundum in sterile distilled water for 1 min before treating with the disarmed _A. tumefaciens_ containing the pBGCP. Transformed cells were cultured on kanamyein-free medium containing 2,4-D and carbenicillin for 2-3 weeks and then on the kanamycin medium for 3-4 months. The developed somatic embryos were transferred to the medium containing NAA, BA and kanamycin and subsequently regenerated into normal-appearing plants. Presence of the PRSV CP gene in the putative transgenic lines was detected by PCR and the expression of the CP was verified by Western blotting. The transgene was nuclearly inherited as revealed by segregation analysis in the backcrossed R1 progeny. From five independent experiments, the average successful rate of transformation was 15.9% of the zygotic embryos treated (52 transgenic somatic embryo clusters out of 327 zygotic embryos treated), about 10-100 times higher than the available methods previously reported. Thus, wounding highly regenerable differentiating tissues by carborundum vortexing provides a simple and efficient way for papaya transformation mediated by Agrobacterium.