Effects Of Seed Priming By N-acetyl-L-cysteine And Melatonin On Seed Germination,Seedling Growth And Gene Expression Of Hybrid Rice Under Low Temperature Stress

Rice is native to tropical and subtropical regions and is susceptible to low temperature stress during the early stages of emergence and seedling development.In the middle and lower region of the Yangtze River and southern China,the"cold spring"is prone to occur in March-April,which will bring serious harm to seed production.Therefore,the effects of seed priming with N-acetyl-L-cysteine（NAC）and melatonin（MT）on cold resistance of rice seeds were studied,using hybrid rice,cv.Longxiangyou130 and Zhuliangyou06 as materials.The germination of seeds subjected to low-temperature stress（15℃）for 11d,then recovery growth（25℃）for 3d,seedling growth,the activity of antioxidant enzyme,the content of glutathione and proline and the expression of genes related to cold resistance were measured.This study provided theoretical basis and practical technology for improving cold resistance of rice seeds.The main results were as follows:1.Effects of priming with different concentrations of NAC on seed germination,seedling growth and gene expression of two cultivars of hybrid rice were studied under low temperature stress.The results showed that,the optimal priming concentrations for Longxiangyou130 and Zhuliangyou06 seeds were 0.1mM and 0.25mM.As compared with the unprimed control,Longxiangyou130 seeds primed with 0.1mM NAC and Zhuliangyou06 seeds primed with 0.25mM NAC,the germination percentage was significantly increased 33.2%and 7.2%after cold stress（11d）;after recovery growth for 3d,the average germination time was significantly shortened by 1.9d and 1.2d,the germination index was significantly increased by 1.2 and 1.5,and the vigour index was significantly increased by 9.4 and 8.5,respectively.Meanwhile,after cold stress（11d）and recovery growth（3d）,the activity of superoxide dismutase（SOD）,peroxidase（POD）,catalase（CAT）,ascorbate peroxidase（APX）,the content of glutathione（GSH）and proline（Pro）in seedling leaves were significantly increased,and the contents of superoxide anion(O₂^·-),hydrogen peroxide（H₂O₂）and malondialdehyde（MDA）were significantly reduced.After priming with NAC,the expression of genes that related to cold signals（OsCPK24,OsP5CS,OsPRP,OsWRKY45）and antioxidant enzymes（OsCATa,OsCATb,OsCATc,OsAPXa,OsAPXb）were significantly up-regulated in both cultivars.It indicated that NAC priming enhanced the antioxidant capacity of seedlings,reducd the damage caused by oxidative stress and increased the cold tolerance of hybrid rice seed.2.Effects of priming with different concentrations of MT on seed germination,seedling growth and gene expression of two cultivars of hybrid rice were studied under low temperature stress.The optimal priming concentrations were 0.2mM in Longxiangyou130 and Zhuliangyou06.As compared with the unprimed control,Longxiangyou130 and Zhuliangyou06 seeds primed with 0.2mM MT,the germination percentage was significantly increased by 18.0%and 16.0%after cold stress（11d）.After recovery growth（3d）,the germination index was significantly increased by 1.5and 1.6,the average germination time was significantly shortened by 1.2d and 1.5d,and the vigour index was significantly increased by 9.7 and 11.6,respectively.Meanwhile,after recovery growth（3d）,the activity of SOD,POD,CAT,APX,the content of GSH in seedling leaves were significantly increased,and the contents of H₂O₂ and MDA were significantly reduced,the expression of genes which related to cold signals and antioxidant enzymes were significantly up-regulated.It suggested that MT priming could reduce MDA content in rice seedlings under low temperature stress by enhancing antioxidant enzyme activity and increasing GSH content to remove excess H₂O₂,thus increased the cold tolerance and improved seedling growth of hybrid rice seed.In conclusion,N-acetyl-L-cysteine and melatonin can be used as priming chemicals.Priming with these two chemicals at appropriate concentrations can promote hybrid rice seed germination and seedlings growth at low temperatures by increasing antioxidant enzyme activity,GSH content and cold-resistant gene expression,enhancing the cold tolerance of hybrid rice seed.