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根据R基因及其调控基因保守序列设计简并性引物 ,对白粉菌接种和未接种处理的一对抗病和感病的小麦 黑麦等位突变易位系TAM10 4R和TAM10 4S总RNA进行RT PCR扩增 ,得到一个诱导表达的cDNA片段。序列分析表明 ,该片段全长 2 4 74bp ,其中含有一个 82 2bp的完整开放阅读框 ,推测其编码一个有 2 73个氨基酸残基、分子量约 31kD的蛋白质分子。蛋白质的氨基酸序列比对显示 ,该蛋白质分子具有C2 HC锌结合motifCX2CX4HX4C结构和锌指domain ,可见克隆的cDNA是一个锌指蛋白基因 ,命名为TaZF。Southern杂交表明 ,TaZF在抗病易位系TAM10 4R的基因组中是多拷贝的。半定量RT PCR分析显示 ,TaZF基因属组成型表达、但受白粉菌诱导表达上调的基因 ,推测其与白粉病菌的侵染过程相关。基因组DNA专化扩增、克隆和测序揭示TaZF基因无内含子。
According to the conserved sequences of R gene and its regulatory genes, degenerate primers were designed and used to detect the total RNA of TAM10 4R and TAM10 4S, a pair of resistant and susceptible wheat rye translocation lines TAM10 4R and TAM10 4S, PCR amplification, to obtain a cDNA fragment induced expression. Sequence analysis showed that the full length of this fragment was 2474bp and contained a complete 82 bp open reading frame. It was deduced that it encoded a protein molecule with 2 73 amino acid residues and a molecular weight of about 31 kD. The amino acid sequence alignment of the protein showed that the protein molecule has a C2 HC zinc binding motifCX2CX4HX4C structure and a zinc finger domain. The cloned cDNA is a zinc finger protein gene named TaZF. Southern blotting showed that TaZF was multicopy in the genome of disease-resistant translocation line TAM104R. Semi-quantitative RT-PCR analysis showed that the TaZF gene is constitutively expressed, but up-regulated by the powdery mildew-induced gene is speculated to be related to the infection process of powdery mildew. Genomic DNA amplification, cloning and sequencing revealed no introns in the TaZF gene.