采用通用引物PCR配合SSCP和RFLP技术检测鱼病病原菌

Universal primer PCR with SSCP and RFLP for identification of fish disease pathogens

  • 摘要: 采用通用引物PCR(UPPCR)、PCR-RFLP、PCR-SSCP技术,研究快速鉴别鱼病病原菌的分子生物学诊断技术。结果发现,采用细菌16S rRNA基因保守区特异性引物,以嗜水气单胞菌、鲁克氏耶尔森菌、鳗弧菌、柱状曲挠杆菌、乙型链球菌、荧光假单胞菌等部分常见鱼病病原菌为对象,可以建立一种UPPCR技术。该技术能在保证实验条件不变的基础上,检出上述所有细菌,并还可检出大肠杆菌和双歧杆菌等非鱼病病原菌。并且认为,该法与SSCP配合即采用UPPCR-SSCP技术能较好地鉴别被检菌而用于鱼病病原菌的快速诊断。

     

    Abstract: A universal primer PCR(UPPCR) , PCR-RFLP and PCR- SSCP had been screened for a molecular biological method that permitted the rapid identif ication of fish pathogens among them. The results showed that a universal primer PCR technology was available with specif ic primers from conserved regions of bacterial 16S ribosomal RNA genes. The bacteria tested included some common causative agents of f ish diseases such as Aeromonas hydrophila , Yersinia ruckeri , Vibrio anguillarum, Flexibacter columnaris, beta-Streptococcus, Pseudomonas f luorescens. The approach allowed the bacteria above and Escherichia coli and Bif idobacterium catenulatum detectable w ithout any alterations of the experiment condit ions. However, this study also found that the identification of species of bacteria tested depended on the combination of UPPCR and SSCP( PPCR-SSCP) , which was better than that of UPPCR and RFLP( UPPCR-RFLP) and made rapid diagno sis of fish disease pathogens possible.

     

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