Agrobacterium-mediated transformation of cucumber (Cucumis sativus L.) using a sense mitogen-activated protein kinase gene (CsNMAPK) View Full Text


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Article Info

DATE

2012-11-30

AUTHORS

Jing Wang, Shoujie Zhang, Xiao Wang, Lina Wang, Huini Xu, Xiufeng Wang, Qinghua Shi, Min Wei, Fengjuan Yang

ABSTRACT

The objective of this study was to develop an efficient regeneration and transformation protocol for cucumber (Cucumis sativus L.). Cotyledonary node explants showed a very high regeneration frequency (94.3 %) when incubated on Murashige and Skoog medium (MS) supplemented with 1.5 mg l−1 6-benzyladenine (BA). An Agrobacterium-mediated transformation methodology for cucumber was developed using this protocol. A construct containing a sense mitogen-activated protein kinase gene (CsNMAPK) was used to transform cucumber cotyledonary nodes. Adventitious shoots were obtained in selection media containing 150 mg l−1 kanamycin and 250 mg l−1 carbenicillin 4 weeks after bacterial inoculation. A total of twelve transgenic plants were confirmed using PCR-specific analysis. The transgene copy number of four transgenic plants (random selection) was found to be a single gene copy using Southern blot analysis and the expression level of the CsNMAPK gene in the transgenic plants was significantly higher than in the wild-type plants. The transgenic rate was approximately 4.8 %. More... »

PAGES

269-277

References to SciGraph publications

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  • 2005-10-25. Improved cotyledonary node method using an alternative explant derived from mature seed for efficient Agrobacterium-mediated soybean transformation in PLANT CELL REPORTS
  • 1990-09. Transformation of Cucumis sativus tissue by Agrobacterium tumefaciens and the regeneration of transformed plants in PLANT CELL REPORTS
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    24 schema:description The objective of this study was to develop an efficient regeneration and transformation protocol for cucumber (Cucumis sativus L.). Cotyledonary node explants showed a very high regeneration frequency (94.3 %) when incubated on Murashige and Skoog medium (MS) supplemented with 1.5 mg l−1 6-benzyladenine (BA). An Agrobacterium-mediated transformation methodology for cucumber was developed using this protocol. A construct containing a sense mitogen-activated protein kinase gene (CsNMAPK) was used to transform cucumber cotyledonary nodes. Adventitious shoots were obtained in selection media containing 150 mg l−1 kanamycin and 250 mg l−1 carbenicillin 4 weeks after bacterial inoculation. A total of twelve transgenic plants were confirmed using PCR-specific analysis. The transgene copy number of four transgenic plants (random selection) was found to be a single gene copy using Southern blot analysis and the expression level of the CsNMAPK gene in the transgenic plants was significantly higher than in the wild-type plants. The transgenic rate was approximately 4.8 %.
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