Cloning Of Gene Related To Fertility Restoration Of C-cytoplasmic Male Sterility In Maize

The use of cytoplasmic male sterility (CMS) is an important approach in the utilization of heterosis. Maize (Zea mays L.) is the first crop that uses cytoplasmic male sterility for hybrid seed production. Since it can reduce the cost and improve the purity of hybrid seed, so the cytoplasmic male sterility (CMS) has become a main type in seed production. However, the genetic of fertility restoration in C-cytoplasmic male sterility is very complicated, and its restoration mechanism is not clear, then it becomes a main restrictive factor in using cytoplasmic male sterility in hybrid seed production. In this study, a related gene of fertility restoration for the CMS-C in maize was cloned by means of transposon tagging method, and its function of the candidate gene was preliminary analyzed. The main results showed in the hereinafter paragraphs.Five mutants of the related fertility restoration were selected in a 29,750 F1 progeny plants between 87-1(Rf4Rf4) and Mu7, and the cytoplasmic type was confirmed by means of three special primers of different cytoplasmic types in maize. The special amplification band was selected using an improvement approach named Mu-AFLP method between two parents and the mutants, an about 750bp AFLP band was found when using a nested Mu7-specific primer Bglâ…¡-2 and a selective primer M1.A Mu7-flanking fragment was obtained after sequenced, which contained partial sequences of Mu transposon and an unknown sequence.According to the sequence of the candidate gene, the PCR primers were designed and a 209bp cDNA was obtained from the cDNA coming from the floret differentiation stage of tassel, by mean of 3'rapid amplification of cDNA end (RACE) method.Using the results of 3'RACE and electronic splicing in Genbank,a 955bp cDNA sequence was obtained; it contained a 321bp open reading frame which encoded 106 amino acid. The full length of the candidate gene was consisted of 2403 bp DNA sequence, contained three introns and four extrons. The results of functional domain analysis showed that the protein was a basic transcription factor TFIIA. It has homologous with the TFIIA gene of Oryza sativa and Arabidopsis thalian, and it has close relationships with the Oryza sativa. Transcription factor TFIIA was a nuclear protein involving in the RNA polymerase II-dependent DNAtranscription, and it was one of several basic transcription factors that were required for all transcription events using RNA polymerase II.The previous reports have indicated that the CREM gene could be affected by the TFIIA in animals, which took an important place in the development of male germinal cells, and TFIIA could indirect regulate the development of maturity at different stages of spermiogenesis through regulating the key gene CREM. So we can suppose that the candidate gene, TFIIA, perhaps has some function on pollen development in maize.