梨ACC氧化酶基因(ACO)的片段克隆及其RNAi载体构建

Cloning of ACC oxidase gene(ACO)of pear (Pyrus pyrifolia) and construction of its corresponding RNAi vectors

以若光梨成熟果实为材料,提取总RNA,在ACC氧化酶基因(ACO)cDNA序列同源性较高区域设计一对特异引物,利用RT-PCR克隆了ACC氧化酶(ACO)基因片段。将ACO反义基因、与副球菌中类胡罗卜素合成有关的间隔基因YYT和ACO正义基因3个片段串联在一起,经鉴定后,插入到植物表达载体中,构建成能够表达ACC氧化酶基因的双链RNA的植物载体pYF028,为耐贮砂梨的遗传转化创造条件。

The total RNA was isolated from the ripe fruit of Wakahikari pear cuhivar （Pyrus pyrifolia NaKai cv. Wakahikari）. A pair of special primers was designed according to the high homology region of A CC oxidase gene. The antisense and sense A CC oxidase （ACO） gene were obtained by RT-PCR and tandem ligated with the gene YYT as internal sequence, which is relative to carotenoid synthesization in subcocci. The fusion gene was inserted into plant vector to construct the recombinant plasmid pYF028, which could express dsRNA ofA CC oxidase gene. In such way it provided the possibility of transformation to improve the storage life of pear cuhivars.