根据已报道的甘薯G病毒（Sweet potato virus G,SPVG）外壳蛋白基因（coat protein,CP）序列,设计和合成了一对特异引物,采用反转录-聚合酶链式反应（Reverse transcription-polymerase chain reaction,RT-PCR）技术,从甘薯病叶总RNA中克隆到了SPVG-CPSC1和SPVG-CPSC2两个约700bp的cDNA片段.该片段与pMD 18-T载体连接后转化Escherichia coli JM109,对筛选到的阳性克隆进行序列测定与分析结果表明,该片段与已报道的甘薯G病毒外壳蛋白基因具有较高的同源性,证实获得的DNA片段是SPVG的外壳蛋白基因.两个克隆片段与埃及分离物EgyPt 1的序列同源性最高,核苷酸同源性分别为98.7%和89.1%,对应推导的氨基酸同源性分别为98.7%和94.8%,与中国分离物CH2的序列同源性最低,核苷酸同源性分别为87.4%和87.0%,对应推导的氨基酸同源性分别为96.6%和94.8%,而2个克隆片段之间的核苷酸以及对应推导的氨基酸序列同源性分别为89.5%和96.1%,表明SPVG具有一定的多态性.
Two cDNA fragments （SPVG-CPSC1, SPVG-CPSC2） with about 700bp were cloned from sweet potato （Ipomoea batats （L.） Lam.） which were suspiciously infected with sweet potato virus by reverse transcription-polymerase chain reaction （RT-PCR） method using a pair of specific primers which were designed based on the conserved region of the coat protein （CP） genes of sweet potato virus G （SPVG）. PCR product was cloned into pMD18-T vector, the linked DNA was transformed into Escherichia coli JM109 and the positive clones were sequenced and analyzed. The results showed that the two cloned fragments were highly identical with known CP genes of SPVG. There was the highest identification between the Egypt 1 isolate and the two clones, with homology of the nucleotide and deduced amino acid sequences of 98.7% and 98.7% for SPVG-CPSC1, 89.1% and 94.8% for SPVG-PSC2, respectively and the lowest identification between the China isolate CH2 and the two clones, with homology of the nucleotide and deduced amino acid sequences of 87.4% and 96.6% for SPVG-CPSC1, 87.0% and 94.8% for SPVG- CPSC2, respectively. While the homology of nucleotide and deduced amino acid sequences between two cloned fragments were 89.5% and 96.1%, respectively, which indicated that the CP gene of SPVG had some polymorphism.
Chinese Agricultural Science Bulletin
Sweetpotato virus G （SPVG）, Coat Protein （CP）, Reverse transcription-polymerase reaction （RT-PCR）, Gene Cloning