关键词: 苎麻, 细胞质雄性不育, CoxⅡ基因, atpA基因, RT-PCR

Abstract:

【Objective】This study aimed at probing into the correlation of mitochondrial genes CoxⅡ and atpA with the cytoplasmic male sterility (CMS) in ramie [Boehmeria nivea (L.) Gaud.]. 【Method】The CMS related CoxⅡ and aptA gene fragments were cloned from the mitochondrial DNA of ramie CMS line and its maintainer and restorer lines (fertile lines) by using PCR and degenerated primer strategy. The primers were designed according to the reserved sequences in the encoding region of mitochondrial genes CoxⅡ and atpA of some dicotyledons from GenBank. The complete CoxⅡ and atpA genes were obtained by means of amplifying the 3′ and 5′ end unknown sequences of these gene fragments using DNA Walking method. Based on the complete gene sequences and gene expression patterns (RT-PCR), the differences of these two genes between ramie CMS line and the fertile lines were analyzed. 【Result】The amplified CoxⅡ and atpA gene fragments showed 95% and 97% homology with the analogous genes from some dicotyledons. The complete CoxⅡ and atpA genes each had an intact open reading frame. The CoxⅡ gene showed no difference between ramie CMS line and fertile lines at the levels of mtDNA sequence, transcription and protein. However, compared to the fertile lines, atpA gene from CMS line showed obvious difference at the levels of mtDNA, amino acid sequence and the secondary structure of protein. And an abnormal low expression of atpA gene in CMS line at the budding stage and late-blooming stage was revealed by RT-PCR analysis. 【Conclusion】The correlation of CoxⅡ gene with ramie CMS could be excluded, but the result suggested a close relationship of ramie CMS with the variation in DNA sequence and / or abnormality in expression of atpA gene in CMS line.

Key words: ramie [Boehmeria nivea (L.) Gaud.], cytoplasmic male sterility (CMS), CoxⅡ gene, atpA gene, RT-PCR