Studies On Raphanobrassica Cytoplasmic Male Sterility Line NRO4270A In Brassica Napus L.

Rapeseed is one of the most important oil crops worldwide. Heterosis has been widely used to increase the yield in rapeseed. Cytoplasmic male sterility(CMS) is one of the important pollination control systems for heterosis utilization recently. Currently,70% of rapeseed hybrid varieties are breeded through cytoplasmic male sterility in China. Pol CMS is still most extensively utilized. Pol CMS easily produces trace pollen, which are influenced by temperature. Meanwhile, the sterile cytoplasm simplification is also a serious problem in rapeseed production. Therefore, it is significant to screen or create stable CMS lines to improve the production of rapeseed hybrid. NRO4270A CMS was found in the distant hybridization offsprings of Raphanus and Brassica by Prof. Wu Jiangsheng. It is a stable cytoplasmic male sterility insensitive to tempature. This research investigated flower morphology, anther development characteristics, genetic classification and molecular feature of NRO4270A CMS through field investigation, cytology observation, analysis of restorer and maintainer relationship and molecular level identification, and confirmed that it is a novel type of cytoplasmic male sterility. The main results are as follows:1. NRO4270A has normal petals, short stamens, ivory transparent and triangular anthers. NRO4270A was completely sterile in different environments for six generations in three years, and its sterility was stable and not influenced by temperature.2.313 rapeseed varieties and the restorers of Pol CMS, Ogu CMS, Kos CMS and WNJ01A CMS were selected for test-crossing with NRO4270A CMS. The results showed that the restorer and maintainer relationship of NRO4270A CMS was different from the other CMS types.3. The microscopic observation results of anther paraffin section indicated that the abortion of NRO4270A microspores occurred from the tetrad stage to the stage of mononuclear pollen grain. The anthers of NRO4270A CMS were normal from primary sporogenous cell stage to tetrad stage, and the tetrads could form. After the microspores with one nucleus were released, the tapetum became vacuolated and radial expandation, leading to extrude the microspore. There was no deposit of sporopollenin outside microspore ektexine, and vacuolization happened in microspores gradually. At the later stage of microspore development, microspores were plasmolysis obviously. The tapetum and microspores degraded absolutely, which resulted in empty anthers and pollenless traits.4. Molecular assays were carried out compare five different CMS types in Brassica napus. Multiplex PCR distinguished NRO4270A CMS with Pol CMS, Ogu CMS, Kos CMS and WNJ01A CMS. Twelve out of 18 combinations of mitochondrial probe/enzyme used for restriction fragment length polymorphism (RFLP) analysis separated the NRO4270A CMS system from the other four systems. Among these 12 combinations,3 combinations including atpl/EcoR I, atpl/BamHⅠand atpl/HindⅢdistinguished each CMS systems from others. Therefore, they can be used as valuable molecular markers to identify the different cytoplasm type. The results further confirmed that the cytoplasm of NRO4270A CMS is different from Pol, Ogu, Kos and WNJ01A CMS at the molecular level. It is a new type of male sterile cytoplasm lines.