马铃薯Y病毒外壳蛋白基因的克隆及全序列测定

CLONING OF POTATO VIRUS Y COAT PROTEIN GENE AND ITS NUCLEOTIDE SEQUENCE ASSAY

自田间采集的马铃薯病叶中提取马铃薯Y病毒 (PVY)总RNA ,人工合成引物P1、P2 ,通过RT PCR扩增合成PVY cp的cDNA ,并将其克隆到 pGEM TEasy载体上。经限制酶谱分析后进行全序列测定 ,结果表明 :该基因由 80 1个核苷酸组成 ,编码 2 67个氨基酸 ,与文献报道的 3个PVY cp基因相比同源性均在 90 %以上 ,且其编码的氨基酸同源性均在 94 4 %以上。说明PVY

Total RNA of potato virus Y (PVY) was isolated from diseased potato leaves collected in field and PVY coat protein (PVY cp) cDNA was synthesized by reverse transcription polymerase chain reaction (RT PCR). Specific PCR product was cloned into pGEM T Easy Vector and the recombinant plasmid (pGEM pvy) contained PVY cp gene was characterized with restriction endonuclease mapping and nucleotide sequencing. The result showed that the PVY cp gene contains 801 nucleotides encoding 267 amino acids. The similarities of nucleotide sequences and amino acid sequences between cloned PVY cp gene and other three published ones were above 90% and 94 4%, suggesting that PVY cp gene is very conservative.