CAJ | 학술논문

目的：建立一种简洁稳定、特异灵敏的检测泰泽病原体（ Tyzzer ’ s organism ）的反向斑点杂交方法（reverse dot blot， RDB）。方法根据泰泽病原体16S rDNA保守基因组序列设计引物和特异性探针，上游引物用生物素标记，进行PCR扩增，建立反向斑点杂交方法，用此法进行了特异性和灵敏度实验，同时，用RDB、ELISA和IFA对41只小鼠、38只大鼠进行了检测。结果 RDB特异性强，最低检测限为4.5 ng／μL。对79例实验动物的检测中，与ELISA检测结果一致性为100％，阳性率是7.59％（6／79）；与IFA一致性为92.4％（73／79），IFA阳性率是0％。结论建立了PCR扩增与分子杂交相结合的准确、灵敏、特异的泰泽病原体反向斑点杂交检测方法。
목적：건립일충간길은정、특이령민적검측태택병원체（ Tyzzer ’ s organism ）적반향반점잡교방법（reverse dot blot， RDB）。방법근거태택병원체16S rDNA보수기인조서렬설계인물화특이성탐침，상유인물용생물소표기，진행PCR확증，건립반향반점잡교방법，용차법진행료특이성화령민도실험，동시，용RDB、ELISA화IFA대41지소서、38지대서진행료검측。결과 RDB특이성강，최저검측한위4.5 ng／μL。대79례실험동물적검측중，여ELISA검측결과일치성위100％，양성솔시7.59％（6／79）；여IFA일치성위92.4％（73／79），IFA양성솔시0％。결론건립료PCR확증여분자잡교상결합적준학、령민、특이적태택병원체반향반점잡교검측방법。
Objective To establish a simple, stable, specific and sensitive method for detection of Tyzzer ’ s or-ganism by reverse dot blotting ( RDB) .Methods Primers and specific probes were designed according to the conservative sequence of Tyzzer 16S rDNA.The forward primer was labeled with biotin .The reverse dot blotting method was established followed by PCR amplification .The specificity and sensitivity of this method were determined .Next, 41 mice and 38 rats were examined by RDB , ELISA and IFA .Results The RDB method showed a high specificity , and in the testing of the 79 laboratory animals , its limit of detection was 4.5 ng/μL.Compared the results of ELISA and IFA , its consistence with ELISA was 100%and the positive rate was 7.59%(6/79), the consistence with IFA was 92.4%(73/79), and the posi-tive rate was 0%.Conclusions An accurate, sensitive and specific method in combination with PCR and RDB in detection of Tyzzer’ s organism is established in this study .