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Efficient transient expression and transformation of PEG-mediated gene uptake into mesophyll protoplasts of pepper (Capsicum annuum L.)

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Abstract

PEG-mediated transformation was used for gene delivery and evaluation of various parameters affecting the transient expression of a gene for ß-glucuronidase (gus) in mesophyll protoplasts of Capsicum annuum. Transient expression was found to be dependent on PEG concentration and exposure time of plasmid DNA to protoplasts as well as the amount of plasmid DNA. Maximum GUS activity was obtained when protoplasts were applied to 40% concentration and molecular weight was 6,000 of PEG solution with 30 min of exposure time. Protoplasts of pepper were transformed with a vector, pCAMBIA::Ac, which contained a pCAMBIA1302 T-DNA vector carrying a maize transposable element, Ac (activator), a selection marker HPT (hygromycin phosphotransferase), and a GFP-coding region driven by the 35S promoter in the presence of PEG. Approximately 30% of the protoplasts expressed GFP. Visibly transformed colonies were obtained from protoplasts after 2 months of culture and GFP was expressed. Southern hybridization confirmed the presence of Ac in the pepper genome.

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Abbreviations

PEG:

polyethylene glycol

PVP:

polyvinylpyrrolidone

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Acknowledgements

This work was supported by a grant from Plant Diversity Research Center (21st Century Frontier Research Program. Project No. PF003202-05), by a grant from KOSEF to the Environmental Biotechnology National Core Research Center (grant #: R15-2003-012-02003-0) and by a grant from the Ministry of Agriculture and Forestry (grant no. 203069-032-2-CG000).

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Authors and Affiliations

  1. Division of Applied Life Science (BK21) and Environmental Biotechnology National Core Research Center, Gyeongsang National University, Jinju, 660-701, Korea

    Joo Mi Jeon, Nam Young Ahn, Bo Hwa Son, Cha Young Kim, Chang-deok Han & Sung-Ho Lee

  2. Department of Microbiology, College of Natural Science, Pukyong National University, Busan, 608-737, Korea

    Gun-Do Kim

  3. Department of Microbiological Engineering, Jinju National University, Jinju, 660-758, Korea

    Sang Wan Gal

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  1. Joo Mi Jeon
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  2. Nam Young Ahn
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  4. Cha Young Kim
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  8. Sung-Ho Lee
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Correspondence to Sung-Ho Lee.

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Jeon, J.M., Ahn, N.Y., Son, B.H. et al. Efficient transient expression and transformation of PEG-mediated gene uptake into mesophyll protoplasts of pepper (Capsicum annuum L.). Plant Cell Tiss Organ Cult 88, 225–232 (2007). https://doi.org/10.1007/s11240-006-9194-z

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  • DOI: https://doi.org/10.1007/s11240-006-9194-z

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