[Objective] This study aimed to develop a quantitative competitive assay for detecting Mycoplasma hyopneumoniae in culture. [Method] One pair of Mhp-specific primers was designed for detecting Mhp in culture. Another ...
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[Objective] This study aimed to develop a quantitative competitive assay for detecting Mycoplasma hyopneumoniae in culture. [Method] One pair of Mhp-specific primers was designed for detecting Mhp in culture. Another pair of primers was designed based on the conserved gene sequences of Mycoplasma. The competitive template, which carried the same primer binding site with the target fragment, was constructed using enzyme digestion method. [Result] The logarithm of concentration of competitive template was treated as the abscissa（X-axis）, and the logarithm of corrected optical density ratio between amplification products by competitive template and target template was treated as the ordinate（Y-axis）. Thus the standard curve was drawn, and the regression equation was also obtained. When Y was assigned as 0, the concentration of the competitive plate was calculated, and then the concentration of Mhp was deduced. The logarithms of Color change unit（CCU） were treated as the abscissa（X-axis）, and the copy numbers of Mhp were treated as the ordinate（Y-axis）, so the standard curve was generated. It was found that the copy number of Mhp was highly correlated to CCU. [Conclusion] A quantitative competitive PCR assay was successfully established for the rapid detection of Mhp in culture.
引言副流感病毒3型属副黏病毒科呼吸道病毒属,是人和动物呼吸道重要病原.其中牛副流感病毒3型(bovine parainfluenza virus type 3,BPIV3)是引起牛呼吸道疾病的主要病原,危害巨大[1,2].2013年底本实验室首次从江苏、安徽多地患呼吸道疾...
引言副流感病毒3型属副黏病毒科呼吸道病毒属,是人和动物呼吸道重要病原.其中牛副流感病毒3型(bovine parainfluenza virus type 3,BPIV3)是引起牛呼吸道疾病的主要病原,危害巨大[1,2].2013年底本实验室首次从江苏、安徽多地患呼吸道疾病的山羊中分离鉴定出山羊PIV3,成为该病毒属的新成员[3].HN、F、M、N是病毒的主要结构蛋白,其中糖蛋白HN具有血凝素和神经氨酸酶活性,介导病毒入胞,也是主要保护性抗原[4].本研究合成JS2013分离株HN基因,构建重组杆状病毒,获得重组蛋白,为病毒HN蛋白的功能研究以及疫苗和检测方法的建立提供有力工具.
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