Observation Of Different Male Sterile Plants And Analysis Of Whole Genome Re-sequencing In Brassica Campestris Ssp.Chinensis

Male sterility is a very common natural phenomenon in angiosperm. Male sterility plants have obvious heterosis, and also play an important part in plant breeding. So breeders pay a lot attention to male sterility plants, and it’s one kind of significant material for scientific researchers as well. Many studies about male sterility on morphology, cytology, physiology and biochemistry, genetics and molecular biology had been finished had been conducted. This article used ’Aijiaohuang’ four male sterility plants including genic male sterility AB line (ajhGMS, Bcajh97-01A/B’), and new-found male sterility plants ’Bc123-26’、Bc123-27’ and male sterility plant in ’Bc123-75’ as experimental material, found abortion stage by morphological, cytological and ultrastructural observation firstly, and then sequenced ’Bcajh97-01A’ and ’Bcajh97-01B’ by whole genome re-sequencing. The sequencing results and data were analyzed, so as to find the mutant gene. The achievement reached was showed as follow:(1) Morphological differences were found in different original male sterile plants through morphological observation. No obvious difference were found in the sepal length, petal length, long anther length, short anther length, long filament length, short filament length and pistil length.between ’Bc123-26’、 ’Bc123-27’.The ’Bc123-15’ male sterility plant was the same. And the petal of ’Bc123-75’ male sterility was more crumpled than the fertility plant. In addition, anther adhesion phenomenon showed in early florescence stage in ’Bc123-26’and ’Bc123-27’.(2) Differences showed in buds of different sizes by cytological observation of male sterility plants from different sources. Scanning electron microscopy (SEM) observation found certain percentage malformed pollens, and the percentage of ’Bc123-27’ was higher than that of ’Be123-27’ in early florescence stage; In later florescence stage, malformed pollens were also found, but the percentage of ’Bc123-27’ was lower; Alexander staining result of pollens indicated no obvious differences between ’Bc123-26’ and ’Bc123-27’ in both early and later florescence stage. DAP1 staining result also showed normal cell nuclei in ’Bc123-26’ and ’Bc123-27’ in later florescence stage; Aniline blue staining result showed callose on the surface of pollens of ’Bc123-26’ instead of ’Bc123-27’; Abnormity of pollen development was found in both ’Bc123-26’ and ’Bc123-27’ in early florescence stage by semithin section observation. SEM observation of pollens of ’Bc123-75’ male sterility and fertility plants indicates much more abnormal pollens found in the former; Alexander staining result of pollens showed differences between ’Bc123-75’ male sterility and fertility plants. DAPI staining result revealed higher aberration rate in male sterility; Aniline blue staining result exhibited callose on the pollens surface of ’Bc123-75’ male sterility while nothing in fertility plants; Semithin section observation found abnormity appeared in mononucleus phase. Pollens of ’Bcajh97-01A’ were 100% malformed, and most pollens linked together; Alexander staining verified pollen viability were 0%.(3) Transmission electron microscope observation of ’Bcajh97-01A’ found abnormity in pollen development. In early mononucleus stage, microspore stage showed evident abnormity and hyperchromatism to see the organelles, and without exine, too. After then, a circle of particles was around the microscope instead of exine. Finally, most pollens degraded, and the left ones adhered together.(4) Many SNPs and Indels between ’Bcajh97-01A’ and ’Bcajh97-01B’ were sequenced by whole genome re-sequencing. These loci were further analyzed to make sure the position(CDS, Non-CDS or Intergenic) of every locus. Finally,90 genes were confirmed as differential genes.(5) Ninty differential genes were homologous cloned and sequenced. The results were not total identicalto the whole genome re-sequencing results.