关键词: 2mG₂-epsps, 18-599R, 农杆菌介导法, 幼胚

Abstract: [Objective] In this study, a new glyphosate-resistant gene 2mG₂-epsps was transformed into immature embryos of maize inbred line 18-599 R by Agrobacterium tumefaciens mediated to obtain transgenic plants with significant resistance to glyphosate. [Method] 2mG₂-epsps gene was introduced into 18-599 R immature corn embryos by Agrobacterium mediated, transformation system was optimized by control the pretreatment conditions, bacterial concentration×immersion time and concentration of glyphosate filter setting process. The 2mG₂-epsps gene had been expressed in transgenic plant by PCR, real time PCR and spraying glyphosate exogenous gene identification. [Results] Heat shock treatment didn't improve the confrontational callus rate, and centrifugation reduced the rate of resistance callus. When the concentration of Agrobacteriumin the inoculation medium was adjusted to OD₆₀₀=0.6, and the infection time was 10 min, the average rate of resistance callus was up to 37.33%. The ideal concentration of Glyphosate was 2.0 mmol/L. Methods using PCR of 46 transgenic plants, 5 positive transgenic individuals were obtained, with a positive frequency of 10.87%. Real-time PCR showed that the foreign gene was expressed in roots, stems and leaves and inherited to next generation stably. And the T₂ generation transgenic plants showed tolerance to glyphosate. [Conclusion] 2mG₂-epsps gene was successfully imported into 18-599 R genome, and gained significant glyphosate resistance.

Key words: maize, 2mG₂-epsps, 18-599R, Agrobacterium tumefaciens, immature embryos