Molecular Genetic Mechanism Of Wheat Male Sterile Mutant NWMS1 And Its Application In Population Improvement

Wheat(Triticum aestivum L.)is one of the most important food crops in the world.Artificial wheat male sterile mutant is of great significance for wheat breeding.In this study,wheat line Shengnong 1 was treated with chemical mutagenic agent EMS,and a stable wheat male sterility mutant named NWMS1 was obtained.The mutant showed abnormal anther development and microspore abortion.The research contents mainly included four aspects: The first was to carry out the phenotype identification of mutant NWMS1;The second was to clarify the genetic analysis of the mutant NWMS1;The third was to study the molecular mechanism of pollen abortion in mutant NWMS1;The fourth was to establish wheat population improvement technology system based on wheat male sterile line NWMS1.The main research results were as follows:(1)Phenotypic identification for mutant NWMS1Most of the agronomic traits of the mutant NWMS1 were similar to those of Shengnong 1.Compared with Shengnong 1,the development process of the mutant NWMS1 was delayed by an average of one week.The leaf apex of mutant NWMS1 was elliptical and light yellow in color from the seedling stage,which could be used as a morphological marker for identification of male sterile plants.Obvious pollen grains could be observed in the anther room of mutant NWMS1,but pollen activity and fertility were very low,and male sterility was complete,self-cross seed setting rate was zero.The floral organs of mutant NWMS1 and Shengnong 1 gradually showed differences from the spore stage.The microstructural observation showed that the meiosis process of the mutant NWMS1 was commonly normal,the spermatocytes of the microspore of the mutant NWMS1 gradually degraded and disappeared.The pollen grains of mutant NWMS1 were oval,severe plasmolysis,no starch grains,and some autophagosomes appeared at the edge of the large vacuole.Apoptosis of microspore cells in mutant NWMS1 eventually led to male sterility at the binucleate stage.Therefore,the mutant NWMS1 belongs to the binucleate microspore abortion type,and the pollen abortion of NWMS1 started at the early uninucleate microspore stage.Pollen grain collapse,plasmolysis,and absent starch grains were three typical characteristics of the abnormal microspores.(2)Genetic analysis for mutant NWMS1Male sterility phenotype of mutant NWMS1 was not related to chromosome structural variation.The offspring phenotypic separation ratio of the isolated population was 1:1,which was controlled by dominant nuclear male sterility gene.The mutant NWMS1 was a novel male sterile mutant,and the male sterile phenotype of the mutant NWMS1 was inherited by single gene dominant inheritance.The gene locus of NWMS1 was located at 2BS.The interval length is 15.52 Mb(78111874 bp…94031598 bp),and Traes CS2B01G120200 was predicted to be a candidate gene of NWMS1.(3)Molecular mechanism of pollen abortion for mutant NWMS1The young spikes or anthers of the mutant NWMS1 and Shengnong 1 were selected for RNA-seq at three stages(S1: the early anther development stage;S2: pollen mother cell meiotic stage;S3: binucleate microspore stage).A total of 1402,170 and 7658 DEGs were identified in the S1-NWMS1 vs.WT,S2-NWMS1 vs.WT and S3-NWMS1 vs.WT sample pairs,respectively.Most valid expressed genes and DEGs were related to nutrient transport and metabolism.K-means clustering showed that 9 metabolic pathways were considered as candidate pathways related to male sterility.WGCNA analysis showed that different classification modules were highly correlated with the samples,and the results of WGCNA analysis were similar to the results of K-means clustering analysis.We identified the main biological metabolic pathways involved in the abnormal development of anther,including protein processing in endoplasmic reticulum,starch and sucrose metabolism,lipid metabolism,and plant hormone signaling.The section tissue staining further confirmed that the loss of starch and other sugars was the main factor for the male sterility of NWMS1 mutant.20 key genes related to abnormal anther development were screened out,including SKP1 B,BIP5,KCS11,ADH3,BGLU6 and TIFY10 B.Therefore,the abnormal anther development of mutant NWMS1 involved a complex gene regulatory network.The primary molecular regulatory model of abnormal anther development had been established successfully.(4)Wheat population improvement based on mutant NWMS1The male sterile line NWMS1 had the advantages of adapting to climatic conditions in Henan Province,high yield,immunity to powdery mildew.The mutant NWMS1 had high general combining ability,and the offspring inherited multiple seeds and good fruiting.At present,wheat population improvement technology based on male sterile line NWMS1 had been successfully established,and population improvement based on male sterile line NWMS1 as the initial maternal parent had achieved good results.The grain bulk density of the lines(or lines)in different combinations of population improvement was mostly above 815 g/L,and the grain hardness level included soft grain,hard grain and medium grain.In addition,the yield potential of progeny lines improved by population improvement was significantly increased by 6.5-8.9%,with an average of 7.8%.Some excellent fertile individual plants had been selected from the offspring of the recurrent population improvement,and entered the identification procedure of line selection,which provided lines for selection and approval of new varieties.