Phenotypic Characterization And Gene Mapping Of Maize Ms2 Male Sterile Mutant

Maize(Zea mays L.) is one of the most important staple crops in the world and the increase of corn grain yields mainly relys on the hybrid vigor. Male sterility is an important agronomic character for the heterosis utility. Cloning of maize genetic male sterile genes not only contributes to the understanding of the molecular mechanism of anther development, but also provides theoretical support for the production of maize hybrid seeds. In this study, we investigated the phenotype of maize ms2 mutant and the stage of pollen abortion. The RNA extrated from anthers at different development stages were used for RNA-seq and the transcripts change was analyzed. We also constructed segretion population and mapped MS2 gene by the map-based cloning method. The main results were as follows:1. At the late stage of anther development, the ms2 mutant exhibits wizened and shrunk anthers which could not produce mature pollen grains. The investigation of the cytological characters of ms2 mutant showed that the abortion of microspores in sterile anthers initiated from young microspore stage. The microspores were degraded completely at the late 1-nucleate stage, leaving debris in the locule. In addition, the ms2 anther shows a defective pollen exine due to that the sporopollenin precursors could not deposite on the prim-exine when the microspores released from tetrad. We also observed that the tapetum of ms2 mutant degraded abnormally and the prim-ubisch bodies were destroyed at late 1-nucleate stage. Moreover, no waxes and cutin monomers were observed on the exo-suface of ms2 anther wall. It is assumed that the function abortion of MS2 gene disturbed the development of tapetum which is responsible for secreting and transporting sporopollenin and lopophillic molecules to the microspore surface and anther wall.2. We performed the transcriptome profiling of wild type and ms2 mutant anthers which grew to 1.5mm, 2.5mm and 3.5mm, respectively. DEGs(differentially expressed genes) analysis revealed that the number of DEGs increased dramatically from 1.5mm stage to 3.5mm stage and most of the DEGs were down-regulated at 3.5mm stage. Subsequently, Mapman was used to analyze the metabolic pathways which DEGs were involved in. The results also showed that the differentially regulated transcripts were enriched in some metabolism pathways, i.e. lipid metabolism, cell-wall-related process, secondary metabolism and lightreaction.3. Previous studies revealed that the MS2 locus is located on a 400 kb region of the long arm of chromosome 9 between TIDP8796 and IDP523. We constructed two segregation populations by crossing ms2 mutant with inbred lines B73 and PH4 CV, respectively. Genetic analysis suggested that the male sterile phenotype is caused by a single recessive mutation. The MS2 gene was mapped on a 6 Mb region of the chromosome 9 between markers InDel8 and InDel10 using 1344 individual male sterile plants in the F2 segregation population of ms2×PH4CV. In addition, 576 individual male sterile plants from F2 segregation population of ms2×B73 were used to map the MS2 locus on a 3 Mb region of the chromosome 9 between markers InDel12 and InDel15. From the above mapping results, we deduced that MS2 gene is located on the long arm of chromosome 9 in a 2.2Mb region between makers InDel8 and InDel15. Subsequently, we constructed a lager segregation population for fine mapping the candidate gene based on the above results. The MS2 gene was mapped on a 380 kb region of the chromosome 9 between markers InDel25 and SNP1 using 4032 individual male sterile plants in the F3 segregation population of ms2×B73.