RNA Editing Of Mitochondrial Functional Genes In Maize Cytoplasmic Male Sterile Lines And Their Maintainer Lines

Maize (Zea mays L.) is one of the plants which utilized heterosis earliest in the world. As to make full use of maize cytoplasmic male sterile lines in heterosis utility, it has an important sense to elucidate the mechanism of cytoplasmic male-sterility (CMS) theoretically. Researchers have made studies associated with CMS in several aspects, such as mitochondrial DNA diversity, plasmid-like, chimeric gene, RNA editing, protein and so on. Therein, RNA editing widely exists in various organisms. It is an important regulating method in CMS mitochondrial gene expression, and according to the research reports, it is supposed that mitochondrial RNA editing is closely related with CMS. Thus researches on CMS mitochondrial RNA editing can help us understand the molecular mechanism of CMS further. Early in 1990s studies on CMS mitochondrial RNA editing had been beginning abroad. But in our country researches on the relationship between RNA editing and CMS in maize has not yet developed. In this thesis, using maize of same nucleus but different cytoplasms and same cytoplasm but different nuclei as the experimental materials, RNA editing of mitochondrial atp6 and cox2 gene in different tissues was analyzed.By direct sequencing, RNA editing of atp6 gene conservative area in two sets of maize tassels of 48-2 and Huang Zao si was analyzed. 18 editing sites were detected out, therein, 4 occurred at the 1st position of codon, 12 at the 2nd position of codon, 2 at the 3rd position of codon. Editing of the 18th site created a stop codon. In different nuclear background materials, except for the 1st editing site of 48-2, editing frequency of sterile cytoplasms was all lower than that of fertile cytoplasms. Occurrence of abnormal edited transcripts of Huang Zao si in sterility was higher than in fertility, but 48-2 was on the contrary. Meanwhile, by clone sequencing, RNA editing of atp6 gene conservative area in 4 cytoplasms' tassels of Huang Zao si was also analyzed. There were 19 editing sites in both N-cytoplasm and S-cytoplasm, 22 in T-cytoplasm, 20 in C-cytoplasm. Their shared editing sites were 18 which were similar to the results of direct sequencing. 1 specific editing site in N-cytoplasm, 2 in C-cytoplasm and 1 in S-cytoplasm all occurred at the 3rd position of codon; 4 specific editing sites in T-cytoplasm, 1 occurred at the 1st position of codon, 2 at the 2nd position of codon, 1 at the 3rd position of codon. Specific editing sites in each cytoplasm appeared in the form of partially editing. By comparing direct and clone sequencing results we concluded that: clone sequencing could detect more editing sites than direct sequencing, and the differential sites of low editing frequency among materials; While, direct sequencing could more exactly reflect the editing frequency and the occurrence of different transcripts.Furthermore, using etiolated seedlings and tassels of sterile lines C48-2, C Huang Zao si and their maintainer lines N48-2, N Huang Zao si as experimental materials, RNA editing of cox2 gene was analyzed by direct sequencing. There were 23 editing sites in materials of 48-2, while there were 22 in materials of Huang Zao si. Their shared editing sites were 22, Therein, 7 occurred at the 1st position of codon, 12 at the 2nd position of codon, 3 at the 3rd position of codon. The 4th site specific to 48-2 occurred at the 3rd position of codon of codon. Cytoplasmic type, tissuey type and nuclear background had little influence on RNA editing of the two genes, but each factor had its own influence tendency on plant editing frequency of the same or different genes. Occurrence of abnormal editing transcripts changed with editing frequency.Synthesizing experimental results, it was concluded as follows: Editing form was C-U reversion; Editing occurring at the 1st or 2nd position of codon would lead to amino acid change except for the 7th editing site of cox2, while occurring at the 3rd position had no effect; Minority of editing sites appeared in the form of partially editing. Differential editing sites were always silent editing sites; RNA editing was inclined to edit bases whose forward was pyrimidine and bases of codons which coded Ser and Pro; RNA editing could improve the predicted protein's hydrophobicity and enhance the conservation among species et.al.In all, the primary results show: most editing frequency of the two genes in sterile lines was higher than in fertile lines and general occurrence of different transcripts decreased specifically in sterile lines. It was supposed that RNA editing of atp6 and cox2 gene had a certain relation with CMS, but it was not the main reason leading to sterility. It might be caused by mitochondrial function degradation when the main factor functioned.