A Novel Laboratory Protocol for Pollen Viability Assessment to Inform Biosafety Evaluation of Transgenic Rice (Oryza sativa L.)

Rice (Oryza sativa L.) is a vital staple crop, and the environmental risk assessment of transgenic varieties is crucial for formulating biosafety policies. Rice pollen grains are spherical, with an average diameter of 40.03 ± 2.75 μm. This study established a standardized protocol for in vitro pollen germination by first optimizing key culture conditions. A single-factor experimental design identified the optimal medium composition as 150 g/L sucrose, 40 mg/L boric acid, 20 mg/L calcium chloride, 10 mg/L monopotassium phosphate, and 10 mg/L magnesium sulfate. The ideal germination temperature was determined to be 31 ± 1 °C, with no germination observed below 16 °C or above 40 °C. Pollen germination rates declined significantly within 5 min post-isolation and ceased completely after 30 min. Building on this optimized protocol, a standardized evaluation method was developed, defining key assessment conditions at temperatures of 25/31/37 °C and post-isolation times of 0/5/15 min. Under these defined conditions, the pollen viability of glyphosate-resistant transgenic rice G2-6 was compared to its non-transgenic recipient ZH11. No significant differences were found at any tested time–temperature combination (p > 0.05). This work establishes a practical and reproducible standard for transgenic rice pollen assessment, offering a scientific basis for evidence-based biosafety regulation and policy-making.