猫杯状病毒2280株反向遗传系统的构建
发布时间：2018-12-13 18:01
【摘要】：为构建猫杯状病毒(FCV)2280株感染性克隆,本研究在FCV 2280株基因组5'末端和3'末端分别引入T7 RNA聚合酶启动子和丁型肝炎核酶(Hdv Rz)序列;并将FCV基因组中唯一的XbaⅠ限制性酶切位点序列进行同义突变作为遗传标记;将基因组分为3段分别进行RT-PCR扩增,通过酶切依次克隆于pOK-12载体中,构建FCV基因组全长cDNA重组质粒。以其为模板经体外转录制备其全长基因组RNA,并在5'末端加帽,转染CRFK细胞,24 h后出现典型的细胞病变。提取出现细胞病变的病毒液的总RNA制备cDNA模板,PCR扩增FCV遗传标记片段并进行XbaⅠ酶切鉴定分析,表明拯救的病毒为FCV重组病毒。而且拯救的病毒生长动力学与亲本病毒无明显差异。本实验为进一步研究FCV的生物学特性和致病机制奠定了基础。
[Abstract]:In order to construct infectious clone of (FCV) 2280 strain of FCV 2280 strain, T7 RNA polymerase promoter and hepatitis D ribozyme (Hdv Rz) sequence were introduced into 5 'terminal and 3' terminal of FCV 2280 genome, respectively. The unique restriction site sequence of Xba 鈪，
本文编号：2376976
[Abstract]:In order to construct infectious clone of (FCV) 2280 strain of FCV 2280 strain, T7 RNA polymerase promoter and hepatitis D ribozyme (Hdv Rz) sequence were introduced into 5 'terminal and 3' terminal of FCV 2280 genome, respectively. The unique restriction site sequence of Xba 鈪，
本文编号：2376976
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