Agrobacterium-mediated transformation of tomato (Lycopersicon esculentum L. cv. Hezuo 908) with improved efficiency
2015
Sun, Sheng | Kang, Xiu-Ping | Xing, Xiao-Juan | Xu, Xiao-Yong | Cheng, Jiao | Zheng, Shao-Wen | Xing, Guo-Ming
We developed an efficient and rapid Agrobacterium-mediated tomato transformation protocol by using cotyledon and hypocotyl as explants. The transgenic nature of the regenerants was confirmed through β-glucuronidase activity staining, polymerase chain reaction (PCR), quantitative real-time PCR (qPCR) analysis and northern blot analysis. In the used protocol, the optimized experimental conditions of the tomato genetic transformation were Agrobacterium liquid concentration, equivalent to optical density (OD₆₀₀) of 1.0, 20 min (cotyledon) or 30 min (hypocotyl) infection time and 2 d of co-culturing period. The optimized medium for shoot induction was Murashige & Skoog (MS) medium + 0.5 mg/L indole-3-acetic acid (IAA) + 0.5 mg/L 6-benzylaminopurine (BAP) for cotyledon and MS + 0.5 mg/L IAA + 2.0 mg/L BAP for hypocotyl. The root induction rate was the highest on the MS medium, supplemented with 0.1 mg/L IAA. The transformation frequency reached 40% per explant. The received results are a base for a larger scale development of transformation procedure.
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