Map-based Cloning Of Male Sterile Gene OsPS11 In Rice

As one of the most important food crops,rice male sterility is not only the basic problem of life science research,but also the basis of crop yield and heterosis utilization.A large number of male sterile genes have been cloned in rice,but the regulatory mechanism and network are not clear.In this study,Nipponbare was mutagenized by EMS,and a homozygous male sterile mutant osps11 was obtained by multi-representative screening.1.The mature anther of mutant osps11 was thinner and smaller than that of Nipponbare,the color was yellowish and transparent,the cuticular structure of the outer wall of anther was abnormal,the number of Urinite on the inner surface of anther decreased and distributed unevenly,no mature pollen grains were produced,and the spikelets were not fruiting,but there was no significant difference in other agronomic characters with Nipponbare.2.The microspore development of the mutant osps11 appeared abnormal micronucleus in the late tetrad stage,the free microspore degenerated,and the degradation of the middle layer and tapetum of the mutant anther wall began to delay in the 9th stage of development.3.The F₁ plants obtained from the cross between mutant osps11 and Nipponbare showed pollen fertility.The genetic analysis of male sterility phenotype in F₂ population showed that the control of male sterility phenotype was controlled by nuclear recessive single gene.4.Using the F₂ population of the mutant osps11 and the wide-compatibility variety Dular to map, 1018 extreme pollen sterile individual plants were screened from the F₂ population,and the gene was finally located on the long arm of the 11th chromosome,between the markers RM27273 and RM224,and the physical distance is 305-kb.5.Using the F₂ backcross population of the mutant osps11/Nipponbare to perform Mut Map analysis, there is a differential gene（Os11g0679700）in the initial mapping interval,and the mutant had a mutation of one base in the second exon of the gene.Resulted in an amino acid substitution.6.The gene was knocked out by CRISPR/Cas9 technology,and the molecular detection and phenotypic observation of the transgenic progenies showed that the homozygous transgenic plants showed no mature pollen grains,the spikelets were not fruiting,and the phenotype was consistent with that of the mutant osps11.We believe that Os11g0679700 is the target gene to control the male sterility phenotype.7.The spatio-temporal expression pattern of Os PS11 gene was studied by quantitative PCR analysis. It was found that the gene was constitutively expressed in various tissues,and the highest expression was found in the anther tissue at the 9th stage of anther development.8.The Os PS11 gene encodes a Glycerol-3-Phosphate Acyltransferase 3（OsGPAT3）,which is located on the endoplasmic reticulum.The defect of anther cuticular layer was caused by controlling the formation of sporophyte in rice,which led to the collapse of anther chamber and sterility.