Identification Of Cold Tolerance And Genome-Wide Association Analysis For Rice Landraces From Different Altitudes In Yunnan

Rice is extremely sensitive to low temperature.Cold damage is one of the most serious natural disaster in rice production,all over the world,around 1/9 of the rice-sown acreage is affected by low temperature.In China,rice yields are reduced by 30–5 billion kg per year due to low temperature and cold damage,which seriously restricts the steady development of rice production.In order to explore the cold-tolerant germplasm and genes that can be effectively used in rice breeding,many scholars have screened queite a few cold-tolerant germplasm resources and genes through the identification and evaluation of cold tolerance of rice germplasm resources and QTL mapping analysis.However,most of these identificationes were confined to a specific growth period in rice growth,and the comprehensive identification of cold tolerance at various growth periods of rice growth and the exploration of genes related to cold tolerance have not been studied comprehensively and systematically.In this study,307 local rice resources from Yunnan Province were used as experimental materials.Through the identification and evaluation of cold tolerance at different growth stages,whole genome resequencing and correlation analysis,the differences in cold tolerance of local rice resources at different altitudes in Yunnan and its correlation with altitudes were clarified.The strong cold-tolerant germplasm at different growth stages was identified,and new genes related to cold tolerance were found.The strong cold-tolerant variety ―Shuainuo‖ and the cold-sensitive variety ―Italynuo‖ were selected to explore the the molecular regulation mechanism of cold tolerance by transcriptome analysis,which is of great significance for molecular breeding of cold tolerance in rice.The research results are as follows:1.Cold tolerance at germination stage,germination stage,seedling stage and booting and flowering stage showed great differences among the tested materials.At low temperature,the germination rate changed to 0-100%,with an average of 38.12%,a standard deviation of 0.346,and a coefficient of variation of 90.77%.The extremely strong cold-resistant materials with grade 1 accounted for 17.92%of the tested materials.The dead seedling rate ranged from 1.67% to 100%,with an average of 69.78%.The standard deviation was 0.293 and the coefficient of variation was 41.96%.The strong cold-resistant materials with grade 3 in cold resistance evaluation accounted for 14.33% of the tested materials.Under low temperature,the mean value of leaf withered degrees was 4.73,the standard deviation was 3.181,and the variation coefficient was 67.31%.The distribution of leaf withered degrees was relatively uniform,and the extremely strong cold-resistant materials with grade 1 accounted for 31.66% of the tested materials.Under natural low temperature at high altitude,the shelling rate(cold tolerance at booting stage)varied from 0% to 100%,with an average of 51.48%,a standard deviation of 0.281,and a coefficient of variation of 54.56%.The excellent cold resistant materials with grade 1 accounted for13.29% of the tested materials.Most of the local rice varieties with strong cold tolerance in the four growth stages are distributed in high-altitude ecological region,while most of the local rice varieties with weak cold tolerance are distributed in the low-altitude ecological region.2.The correlation analysis showed that the cold tolerance of Yunnan local rice varieties at different growth stages was significantly positively correlated with the altitude of their ecological regions,that is,the cold tolerance of Yunnan local rice varieties was closely related to the altitude of their ecological regions.The cold tolerance of local rice varieties in high altitude ecological regions was significantly stronger than that in low altitude ecological regions.The results showed that there was a significant positive correlation between the cold tolerance of each growth period,which indicated that the cold tolerance of each growth period could be used as an indirect index to evaluate the cold tolerance.3.Through the identification and evaluation of cold tolerance at four growing stages,20 accessions of rice landraces with very strong cold tolerance such as Lengshuigu,Cizhonghuanggu,Jiegunuo,Longchuannuo,etc.,which reached grade 1 in both the germination and the seedling stages.9accessions of rice landraces with very strong cold tolerance such as Xiaoheigu,Qitougu,Yunnangu,etc.,which reached grade 1 in both the germination and the booting and flowering stages.18 accessions of rice landraces with very strong cold tolerance such as Bayuenuo,Landigu,Aolenggu,etc.which reached grade 1 in both the seelding and the booting and flowering stages.The very strong cold tolerance germplasm "Aijiaoxi" was screened out,which was 1 grade cold tolerance at the germination stage,the seedling stage,the booting and flowering stage.These excellent germplasmes with very strong cold tolerance are of great value for breeding improvement and new gene discovery for cold tolerance in multiple reproductive periods.4.A total of 67 loci significantly associated with cold tolerance in four growth stages were identified by GWAS analysis.Two loci significantly related to cold tolerance at germination stage were detected.Twenty loci significantly associated with cold tolerance at bud stage were detected.34 loci were found to be significantly associated with cold tolerance at seedling stage.Eleven loci significantly associated with cold tolerance at booting stage were detected.Twelve cloned genes were detected in the four significant association sites of cold tolerance at reproductive stage,which may play a role in cold tolerance.5.Strong cold-tolerant variety ―shuainuo‖ and cold-sensitive variety ―yidalinuo‖ were selected from the test materials,and transcriptome analysis was performed at seedling stage.The results showed that low temperature could cause leaf atrophy,curl,yellowing,slow growth and even death.At the transcriptome level,low temperature stress can cause the changes of plant hormone signal transduction,ribosome composition,photosynthesis,phenylalanine biosynthesis and phenylalanine,tyrosine and tryptophan biosynthesis.WRKY transcription factor plays an important role in the process of biological stress.A novel WRKY transcription factor(novel.1979)was found in the prediction of new genes in this experiment,which may be related to cold tolerance at seedling stage of rice and provide an important reference for subsequent gene function verification.