Isolationing And Translating The Mark Of Mitochondrial Genes Associated With Cytoplasmic Male Sterility In Cotton

Plant cytoplasmic male sterility (CMS) are wide spread in nature. As CMS played a significant role in production praction, it has become a hot research, but its molecular mechanism is still not clear.More research proves that CMS is closely related with the mitochondria, and some CMS-associated mitochondrial genes had been detected in some plants. Although heterosis is very obvious in cotton, the mechanism of CMS of cotton is still at the initial stage comparing to other plants.In order to better reveal the mechanism of CMS, G. harknessii Brandegee S and its maintainer F were used as materials in this research, and the method of Genome walking and RACE (Rapid-amplification of cDNA ends) were performed to analyse the flanking and cDNA sequences in CMS and maintainer, by this method, we got SCAR (Sequence-characterized Amplified Region) marker associated to CMS. and by method of RAPD,we analysed the three phentype materials with near isogenic lines constructed.Here are the main results and conclutions of the research:1. The CMS and its maintainer's flanking sequence of mitochondrial gene atpA were gotten by PCR, including 1678bp of CMS and 1898bp of maintainer,we got atpA gene 11351bp(5766bp of maintai-ner and 5585bp of CMS) including prior period results.2. According to the diversity sequences of gene atpA, we designed the SCAR primer, and performed the PCR using 11 paris of CMS and their maintainers as templates, from many results, primer up-1+s-1 was chosen to identify CMS,primer up-1+F-2 was chosen to identify the maintainer.3. The Sequence of cDNA was cloned by RACE, the results showed they were 3138bp, including CMS is 1582bp and maintainer is 1556bp. The identity of them is 99.6%, but their predict protein coding sequence are different, it may cause CMS.4. Construct the NIL of restorer, there are 3 phenotypes:more pollen, less pollen and CMS; field surveyed the proportion of them is 28:85:106, it is special, we conferred CMS had an impact on the improvement of male gamete.5. According to the knowable primer of RAPD marker and STS marker related to restorer gene, we carried out PCR to restorer line and CMS,the result revealed that there were specific fragments in PCR of UBC111 and UBC679 primer, the result of recovering the polymorphic fragments showed UBC679's fragments are similar to Populus trichocarpa's protein sequence; there were polymorphic fragments in STS-UBC607 in restores line,but there were no fragments in CMS, the primer STS-UBC 679 produces no bands in three materials.