杯状病毒基因组全长cDNA克隆的构建及感染性病毒RNA的体外转录

CONSTRUCTION OF A FULL-LENGTH cDNA CLONE OFCALICIVIRUS GENOME AND TRANSCRIPTION OF RNA

摘要: 目的：构建猫杯状病毒（FCV）基因组全长cDNA克隆，体外转录感染性病毒RNA作为研究人及其他杯状病毒的模型或载体。方法：用RT－PCR法扩增覆盖FCV全基因组的3个首尾重叠的片段，分步插入到表达性载体中，内建全程cDNA克隆，体外转录RNA并转染CrandellReese猫肾（CRFK）细胞。结果：构建成功FCV全长cDNA克隆，体外转录并转染CRFK细胞成功．培养上清对CRFK细胞有感染性，使其出现病变。结论：重组FCV系统将为人及其他杯状病毒的研究提供有效的遗传控制及生物学分析的有效工具。

Abstract: Aim: To construct a full - length cDNA clone of feline calicivirus (FCV) and transcribe infectious RNA from the engineered cDNA, as the model and vector for the researeh of human caIicivirus. MethodS: The clone was construct-ed in a transcription vector by assembly of 3 overlapping PCR fragments, viral RNA was transcribed from the cDNA and tran-fected into Crandell Reese feline kidney (CRFK) cells. Results: The full - length cIone of FCV was constructed. The RNA was transcribed in vitro and transfected into the CRFK cells successfully. The supernatants of culture were infectious to fresh cells and caused the cells ill and cytolysis. Conclusion: The engineered system of FCV should open the way for genetic manipu-lation and biological analysis of the calicivirus genome.