Contracaecum rudolphii姊妹种特异PCR鉴定方法的建立  

The Establishment of Specific PCR Assays for the Identification of Members of the Contracaecum rudolphii Complex

作　　者：何芳[1] 翁亚彪[1] 曹湛[1] 林瑞庆[1] 陈红玲[1] 宋慧群[1] 朱兴全[1] 

机构地区：[1]华南农业大学兽医学院,广州510642

出　　处：《畜牧兽医学报》2005年第9期918-922,共5页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：国家杰出青年科学基金项目(30225033)

摘　　要：参考已测得的C.rudolphii姊妹种(C.rudolphiiA和C.rudolphiiB)及C.septentrionale的核糖体DNA(rDNA)内转录间隔1、2(ITS-1及ITS-2)核苷酸序列,设计、合成了针对C.rudolphiiA、C.rudolphiiB及C.septentrionale的特异性引物HFA(F)、HFA(R)、HFB(F)、HFS(F),通过PCR条件优化和扩增后,它们均能特异地扩增出目的DNA片段,分别是323、321、108 bp,而其它对照样品均未扩增出特异性片段,敏感性试验可检测到最低浓度分别为1.6、21.80、.27 ng/μL。从而初步建立了鉴定C.rudolphii姊妹种的特异PCR方法。该方法特异性强、敏感度较高、重复性好,不仅可用于C.rudolphii姊妹种的分类鉴定,也可用于它们所导致的寄生虫病的诊断和流行病学调查。Species-specific primers for Contracaecum rudolphii A, Contracaecum rudolphii B and Contracaecum septentrionale, namely HFA（F）, HFA（R）, HFB（F） and HFS（F）, were designed based on sequences of the first and second internal transcribed spacers （ITS-1 and ITS-2） of ribosomal DNA. Under optimal amplification conditions, specific rDNA fragments of 323bp, 321bp and 108bp in length were amplified by PCR for C. rudolphii A, C. rudolphii B and C. septentrionale, respectively, while no fragment was amplified from gDNAs from other parasites used as controls. The lowest DNA concentration that the assay could amplify was 1.6, 21.8 and 0.27 ng/μL for C. rudolphii A, C. rudolphii B and C. septentrionale, respectively. These assays could be used for the accurate and rapid identification of members of the C. rudolphii complex, and for the differentiation between C. septentrionale and the C. rudolphii complex. These methods could provide tools for the diagnosis and epidemiological survey of infections these anisakids caused.

关 键 词：Contracaecum rudolphii 特异PCR 特异性 敏感性 PCR方法 鉴定方法 姊妹种 特异性引物 DNA片段 

分 类 号：S513.035.1[农业科学—作物学] TS207.3[轻工技术与工程—食品科学]