Mechanism Study On Radiosensitivity Difference Of Different Japonica Varieties To Carbon-ion Beam Irradiation

Rice(Oryza sativa L.),one of the most important food crops in the world,provides staple foods for most people.It is essential to cultivate new rice varieties with high yield and quality and stress resistance for ensuring food security.In plant mutagenesis breeding,previous studies showed that heavy-ion beam irradiation has unique physical and biological advantages with high linear energy transfer(LET),relative biological effectiveness(RBE),high mutation rate and wide mutation spectrum.The application of heavy-ion beam in rice mutagenesis breeding will expand the genetic diversity of rice,innovate germplasm resources,and better serve ecological agriculture.However,different organisms,organs,tissues,and cells present distinct sensitivity to heavy-ion beam radiation,therefore,diverse mutation processes and receptor damage may occur even under the same irradiation conditions.Overall,exploring the mechanism of radiosensitivity differential in rice may become a critical way,which not only reveals the mechanism of heavy-ion beam mutagenesis but also provides theoretical guidance for the formulation of the optimal irradiation scheme and the application of heavy-ion beam in the breeding of rice and other crops.In this paper,the seeds and seedlings of japonica(Oryza sativa subsp.japonica)were selected to explore the effects of carbon-ion beam irradiation on seed germination and seedling growth,the mechanism of radiosensitivity differences was revealed from the perspective of morphology,cytology,physiology and biochemistry,transcriptomics and metabolomics.The main research results of this paper are as follows:(1)Screen the sensitive and insensitive japonica varieties under carbon-ion beam irradiation.The results showed that carbon-ion beam(0 Gy,80 Gy,160 Gy,80 Me V/u)irradiation had different effects on the germination of 12 japonica varieties.Among12 varieties,irradiation significantly decreased the germination potential of ’Jijing809’ and ’Longdao 18’(LD),the germination potential was 25.83% and 31.67% under80 Gy irradiation,and it was only 3.33% and 5% under 160 Gy irradiation,respectively.Low-dose irradiation hardly affected the germination rate,which was above 80% under 80 Gy irradiation.Except for ’Tonghe 66’(TH),a high dose of irradiation(160 Gy)markedly decreased the germination rate of most varieties.The four varieties with the lowest germination rate were ’Dongdao 4’,’Dongdao 122’,LD,and ’Suijing 18’,which were only 21.67%,22.50%,31.67%,and 31.67%,respectively.TH had the highest germination rate which was as high as 90% after 160 Gy irradiation.The results showed that among the 12 japonica varieties,irradiation had the least inhibitory effect on TH and the most on LD.Therefore,we defined TH as an insensitive variety and LD as a sensitive variety.The dry and wet seeds of LD and TH were selected to explore the influence of water content on radiation sensitivity.The results showed that the germination potential,germination rate,and seedlings survival rate of dry and wet seeds in the two varieties declined sharply with the increase of irradiation dose.And the morphological indexes of TH were always superior to LD under the same irradiation dose.In addition,the negative effect of irradiation on the germination of LD and TH wet seeds was lower than that of dry seeds.However,with the growth of seedlings,the severe internal damage of wet seeds gradually appeared,in the form of a sharp decline in survival rate.After irradiation,wet seeds of LD only had viable seedlings at 80 Gy,and the survival rate was only 5%.But,dry seeds of LD had a certain survival seedlings under each dose,for example,80 Gy irradiation caused a 38.89% survival rate.Similarly,the same dose irradiation resulted in a higher survival rate of seedlings in TH dry seeds than that of wet seeds.It is speculated that abundant reactive oxygen species(ROS)were generated by water radiolysis,and they may participate in wet seeds germination in a short time.However,seeds with high water content present active physiological activity,and ROS will continue to disrupt seed metabolic activities.This may be related to the fact that wet seeds are more sensitive than dry seeds.(2)Study on the radiosensitivity difference of different japonica varieties to carbon-ion beam irradiation.In this experiment,the dry and wet seeds of LD and TH were selected and explored the effects of irradiation on ROS content,antioxidant enzyme activity,and non-enzymatic antioxidant content in their germinating seedlings.The results showed that the ROS and malondialdehyde(MDA)content in germinating seedlings were significantly increased after irradiating the dry and wet seeds of LD and TH,the oxidation level in LD was higher than that of TH,while its antioxidant enzyme activity and non-enzymatic antioxidant content were lower than that of TH.At the same time,compared with the dry seeds of the two varieties,the wet seeds accumulated more ROS,and their antioxidant system was more stronger under the same irradiation dose.It is demonstrated that the ROS and antioxidant system may affect the radiosensitivity difference between TH and LD,and between their dry and wet seeds.(3)Study on the radiosensitivity difference of japonica seedlings to carbon-ion beam irradiation.In this study,LD and TH seedlings were selected to explore the differences in radiosensitivity and physiological indexes of seedlings.The results showed that the stem length,root length,stem weight and root weight of LD were significantly inhibited by irradiation,which were reduced by 27.20%,14.91%,22.93%and 39.95%,respectively,but the root length of TH was only inhibited by 9.76%,and irradiation had no significant negative effect on its stem length and fresh weight.MDA and ROS contents in LD and TH were significantly increased under irradiation,and ROS tissue staining of seedlings also showed similar results.In addition,irradiation also activates antioxidant systems,the activities of CAT and SOD in TH seedlings were higher than that of LD.Irradiation significantly increased the membrane permeability of LD and TH root tips,at 6 h after irradiation,their permeability was increased by 141.05% and 46.65% than the control.At 12 h,the membrane permeability of the LD root tip increased by 101.10%,while TH did not change significantly.The electrical conductivity in LD’s stem was increased by 47.83%and 48.02% at 12 and 48 h,respectively,and it was rose by 17.64% in LD’s root at 12 h,while TH did not increase significantly.Meanwhile,irradiation significantly affected the DNA damage of stems,and the DNA damage degree in LD was more serious than that of TH.The results showed that the stronger antioxidant system in TH seedlings could timely remove excess ROS and maintain redox homeostasis,in addition,it has lower cell membrane damage and DNA damage degree than that of LD,which may be partly responsible for its higher radiation tolerance than LD.(4)Omics analyze the radiosensitivity difference of different japonica seedlings to carbon-ion beam irradiation.In this study,LD and TH seedlings were selected to further explore the mechanism of distinct radiosensitivity by transcriptomic and metabolomic.The integrative analysis showed that there were differentially expressed genes and metabolites in phenylpropanoid biosynthesis,phenylalanine,tyrosine,and tryptophan biosynthesis,linoleic acid metabolism,and alpha-Linolenic acid metabolism pathway,and the distinguish of abiotic stress resistance may be the key factors for the radiosensitivity difference between TH and LD seedlings.In addition,the key genes Os05g0135000,Os03g0434800,Os04g0651000,Os01g0591000,Os11g0644800,Os03g0700400,and Os12g0448900 involved in the above pathways may be the key target genes of irradiation and they may involve in the regulation of japonica radiosensitivity difference.Overall,this study provides new insights and complements partly existing knowledge on the mechanism of radiosensitivity differences among different japonica varieties.