运用基因芯片技术检测三种寄生虫方法的研究

Gene chip assay for detection and identification of three foodborne parasites

目的:利用基因芯片技术建立高效、特异的检测食品中绦虫、弓形虫和旋毛虫的方法。方法:本文选用弓形虫B1基因、旋毛虫SB2基因、绦虫12SrDNA上的可变区设计引物及探针,点样于玻片制成基因芯片。寄生虫DNA经过引物扩增后与芯片上的探针杂交,然后通过扫描图象对结果进行判断。对基因芯片检测的特异性、灵敏性进行了评价,并对模拟污染样本进行了检测,以验证所建立的方法。结果:设计的引物能够扩增三种寄生虫,而且探针的特异性很高。本体系的基因芯片杂交检测敏感度约为15 ng/m l。以食品中旋毛虫为例,可以检测出1条旋毛虫/200 mg。实际检测模拟污染的样本的正确率达到100%。结论:成功制备了检测弓形虫、旋毛虫和绦虫的基因芯片,具有快速、灵敏、特异等特点。

Objective：To develop a accurate detection system based on the promising DNA Chip technology for foodborne parasites. Methods：Choosing the T. gondii B1 gene,T.spiralis SB2 gene, tapeworm 12sDNA variable region, we designed the three primers and probes. Then the probes were immobilized onto certain glass slides that were used as DNA Chip in the experiment. The three kinds of parasites＇DNA were amplified, and the purified amplicons were hybridized to the DNA Chip. Then a fluorescent scanner was used to observe and record the hybridization signals. Using the DNA Chip to detect specificity, sensitivity and stability. The hybridization results were used to evaluate the efficiency of this detection system. Results： The desigened primers could amplify the three kinds of parasite＇DNA, and the probes had high specificity and stability. The detection limit of this system was 15 ng/ml. It can detect one T. spiralis/200 mg. The pathogens in the samples contaminated artificially could be also detected accurately. Conclusion：The genchip technique for detection of T. gondii, T. spiralis and rapeworm has high specificity and sensitivity.