| Rice is an important food crop,more than half of people eat rice as staple food all over the world.At the same time,rice is also a saline and alkaline sensitive crops,saline and alkaline stress has become the main limiting factor in many rice growing areas.There was a significant difference between salt and alkaline resistance of rice,and the influence is greater under alkaline stress.As alkaline resistance is a complex quantitative trait,related studies are few and the QTL contribution rate is low,so fine mapping and cloning is difficult.Therefore,it is necessary to explore and understand the the main effect QTL and genetic mechanism of alkali tolerance in rice,and lay a foundation for the cultivate alkaline resistance rice varieties using molecular markerassisted selection.In this study,the RIL population derived from the cross “DN425×CB10” were used to dentify the alkali-tolerance for two consecutive years.The phenotypic values of granin number per panic,seed setting rate,thousand grain weight,panicle weight per plant and alkaline tolerant coefficients were analyzed by QTL.At the same time,through the identification of phenotypes under many years,the DNA pool was constructed with the thousand grain weight as the target trait,and the correlation of alkali-tolerant QTL interval was carried out with the next generation sequencing technique.By comparing the results of QTL analysis and BSA-seq,the alkali-tolerance candidate genes were mined in the common interval of the two analysis methods,so as to mine the alkali tolerance candidate genes.The main results were as follows:(1)The yield related traits of RIL and parents were decreased in different degrees under the alkali stress.and the decline rate of DN425 stronger than CB10 Granin,it indicates that the alkali resistance of CB10 is stronger than DN425 under alkali stress.Number per panic,seed setting rate,thousand grain weight showed significant or very significant differences between the parents.The range of phenotypic variation of each character is wide,which was in line with the normal distribution.It is suitable to use BSA-seq for QTL mapping analysis.(2)QTL Ici Mapping V4.0 was used to mapping the QTL in 180 RIL population,a total of 13 additive QTLs were detected,which distributed on chromosomes 1,2,3,4,5,8,9 and 11.q GW2 was a main QTL,it was repeatedly detected under alkaline stress for two year s,the contribution rate was 7.77%-23.25%,and synergistic allele was from the paternal CB10.q GN3,q GW2,q GW8,q PW1 and q PW9 were identified as new QTL without any coincidence with previous research results.(3)The BSA-seq method was used to construct thousand grain weight resistance pool for sequencing.The clean reads was 25.48 Gbp,Q30 was 80%,and the average sequencing depth of each sample was 34.17×,the average ratio of sample to reference genome was 97.48%,the average coverage depth was 29×,and the genomic coverage was 98.15 %.Based on the association algorithm of ED and SNP-index,the target interval that a size of 2.89 Mb was mapped on chromosomes 2 and 3,and including 593 candidate genes.(4)The results of QTL mapping and BSA-seq analyzing that the target interva was narrowed to a range of 465 Kb in size of 23.865-24.300 Mb and 25.210-25.240 Mb on chromosome 2.There are 65 genes in this region,and 47 genes were annotated.(5)Through the functional annotation analysis,11 candidate genes related to alkali tolerance were identified,which were LOC_Os02g39750,LOC_Os02g39930,LOC_Os02g39620,LOC_Os02g39884,LOC_Os02g39740,LOC_Os02g39890,LOC_Os02g39764,LOC_Os02g39840,LOC_Os02g39790,LOC_Os02g39960,LOC_Os02g39910 respectively.LOC_Os02g39750 and LOC_Os02g39930 were related to the response of alkali stress and repair function of cell membrane in rice.it can be proposed that LOC_Os02g39750 and LOC_Os02g39930 were correlated with alkali tolerance in rice. |
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