Hybrid rice breeding offers an important strategy to improve rice production, and therefore is crucial for ensuring worldwide food security. Purity of hybrids is a key factor in keeping the high production of hybrid rice. While another important thing is mechanical weeding of hybrid rice, which could greatly release labor from laborious work and cut the production cost. Both of these two problems could be partly resolved by developing glyphosate-tolerant restorer lines. In this study, a novel glyphosate-tolerant gene I.variabilis-EPSPS* was transferred into elite indica restorer line Minghui 86 (MH86) by Agrobacterium-mediated transformation. By Southern blot and inverse PCR analysis, 3 transgenic restorer lines (MY28, MY50, and MY51) with a single copy of I.variabilis-EPSPS* integrated into the intergenic regions were selected from 72 regenerated transgenic plants. The inheritance of I.variabilis-EPSPS* in the 3 transgenic restorer lines was proved to follow Mendelian inheritance by germinating test. In the following generations, according to Southern blot, Northern blot, 3’RACE, and Western blot, I.variabilis-EPSPS* were proved to stably inherit and express in the candidate transgenic restorer lines. At the same time, glyphosate tolerance assay was performed in the field, in which the candidate transgenic restorer lines kept normal agronomic traits under glyphosate treatments with a dosage as high as 8400 g/ha. Therefore, the candidate transgenic restorer lines were highly tolerant to glyphosate. The 3 candidate transgenic restorer lines were respectively hybridized with the male sterile line II-32A, and their hybrids were respectively named as MY28H, MY50H, and MY51H. According to Southern blot and Western blot analysis, I.variabilis-EPSPS* could stably inherit to F₁ hybrids and express in these hybrids, which indicated that the true hybrids could be distinguished by I.variabilis-EPSPS*. In the field test, the transgenic hybrids with I.variabilis-EPSPS* could suffer 3360 g/ha glyphosate without the damage of agronomic traits, which indicated the application potential of these hybrids in direct seeding. Collectively, the transgenic glyphosate-tolerant restore lines developed in this study were highly tolerant to glyphosate, and they had potential to be used in commercial production of hybrid rice.

杂交水稻育种提供了提高水稻产量的重要战略，因此对于确保全球粮食安全至关重要。杂交稻的纯度是保持杂交稻高产的关键因素。杂交稻的机械除草是另一个重要的事情，它可以大大减轻繁琐工作中的劳动并降低生产成本。这两个问题都可以通过开发耐草甘膦的恢复系来部分解决。在这项研究中，一种新颖的耐草甘膦基因I.variabilis-EPSPS *转移至精英籼稻通过农杆菌介导的转化恢复系明恢86（MH86）。通过Southern杂交和反向PCR分析，获得3个转基因恢复系（MY28，MY50和MY51），单拷贝从72个再生的转基因植物中选择整合到基因间区域中的I.variabilis-EPSPS *。通过发芽试验证明3个转基因恢复系中I.variabilis-EPSPS *的遗传遵循孟德尔遗传。在随后的几代中，根据Southern印迹，Northern印迹，3'RACE和Western印迹，I.variabilis-EPSPS *被证明在候选转基因恢复系中稳定地继承和表达。同时，在现场进行了草甘膦耐受性测定，其中候选转基因恢复系在草甘膦处理下以高达8400 g / ha的剂量保持正常的农艺性状。因此，候选转基因恢复系对草甘膦具有高度的耐受性。将3个候选转基因恢复系分别与雄性不育系II-32A杂交，并将其杂种分别命名为MY28H，MY50H和MY51H。根据Southern杂交和Western印迹分析，I.variabilis-EPSPS *可以稳定地遗传到F ₁杂种中并在这些杂种中表达，这表明真正的杂种可以通过杂交来区分。I.variabilis-EPSPS *。在田间试验中，带有I.variabilis-EPSPS *的转基因杂种可遭受3360 g / ha的草甘膦破坏而不会破坏农艺性状，这表明这些杂种在直接播种中的应用潜力。总的来说，这项研究中开发的耐草甘膦转基因恢复系对草甘膦具有高度的耐受性，并且有潜力用于杂交水稻的商业化生产。