HRP直接标记属特异性基因探针检测沙门氏菌的研究

以活化辣根过氧化物酶复合物（ＨＲＰ—ＰＢＱ—ＰＥＩ＋—ＮＨ３＋）直接标记沙门氏菌属特异性ＤＮＡ探针ｐＬＳ２和ｐＬＳ３，探针与靶ＤＮＡ杂交后催化发光底物，经增强型化学发光反应（ＥＣＬ），用普通Ｘ光胶片自显影（ＣＰＤ）检测沙门氏菌。经狭缝杂交（Ｓｌｏｔｂｌｏｔ），该法标记探针均可检测到０．１ｐｇ的纯质粒ＤＮＡ及１０３个未经培养的鼠伤寒沙门氏菌。Ｄｏｔ—ｂｌｏｔ杂交结果证明，ＨＲＰ标记的探针仅与沙门氏菌属细菌杂交，而与试验的其他肠道非沙门氏菌不杂交。本研究表明，ＨＲＰ直接标记基因探针化学发光自显影法检测沙门氏菌，安全、快速、简便，且有高度的敏感性和特异性，是一种有较大应用前景的非放射性标记探针杂交检测方法。

Development of a Test for Detection of Salmonella spp.byDNA Probe Conjugated with Horseradish Peroxidase

The authors have developed a DNA-DNA hybridization test for detection of Salmonella spp.by probe directly conjugated with the activated horseradish peroxi- dase complexes(HRp-PBQ-PEI -NH 3).The binding of the probe to its target was as-sured by the addition of HRP substrates hydrogen peroxide and luminol in the persenceof an enhancer,and the chemiluminescence was recorded by exposure to X-ray film. Dotblot hybridization demonstrated that the HRP-conjugated pLS2, pLS3 probes specificallyhybridized with the DNA isolated from Salmonella spp.Slot blot hybridization showedthat the HRp-conjugated probes were able to detect 0.1 pg of plasmid DNA(pLS2,pLS3)and as little as l03 unincubated Salmonella typhimurium cells. These results sug-gested that detection of Salmonella spp. with the HRp-conjugated gene probe is sensi-tive, specific and has potential diagnositic application in the clinical and/or food hygienelaboratory.