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对棉铃虫Helicoverpa ar migera核型多角体病毒HearSNPV的ORF33基因(ha33)进行克隆和原核表达,hass在E.coli中表达不完全,表达产物的大小为17kDa,小于预测的分子量28.4kDa。用纯化的原核表达产物免疫家兔,制备了多克隆抗体,应用多克隆抗体检测了HearSNPV感染的宿主细胞(HzAMI)中ORF33基因的表达,表达产物的分子量为31kDa。并通过共聚焦荧光显微镜方法,用多克隆抗体检测编码的蛋白在宿主细胞(HzAM1)中的亚细胞定位,发现ha33编码的蛋白存在于宿主细胞的细胞质中,并持续到感染后期。
The ORF33 gene (ha33) of Helicoverpa ar migera nuclear polyhedrosis virus HearSNPV was cloned and prokaryotic expressed. The expression of hass was not complete in E.coli. The size of expressed product was 17kDa, which was less than the predicted molecular weight of 28.4kDa. The purified prokaryotic expression product was used to immunize rabbits to prepare polyclonal antibody. Polyclonal antibody was used to detect the expression of ORF33 in HeLaSNPV infected host cells (HzAMI). The molecular weight of the product was 31 kDa. The subcellular localization of the encoded protein in the host cell (HzAM1) was detected by confocal fluorescence microscopy with the polyclonal antibody. The ha33-encoded protein was found in the cytoplasm of the host cell and lasted until later stages of infection.