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RAPD分析在Hp菌株鉴定中的应用 |
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王蔚虹 阎赞华 胡伏莲
【摘要】 目的 建立一种快速准确地鉴定不同幽门螺杆菌(Hp)菌株的RAPD分析方法,并评价它在区分Hp复发和再感染中的应用价值。方法 用苯酚-氯仿方法提取Hp DNA,以一个10nt的寡核苷酸引物,建立Hp PCR扩增体系,所得PCR产物以2％琼脂糖凝胶电泳分析(RAPD分析)。同一菌株验证方法的重复性，对4例十二指肠溃疡伴Hp感染者三联治疗前后及根除后一年内再现的Hp菌株进行RAPD分析,鉴别复发和再感染。结果 (1)同一Hp菌株于不同时间进行的2次RAPD分析,所得的PCR产物完全一致,说明该方法具有良好的可重复性。(2)50株不同的Hp菌株各产生不同的RAPD图谱,可按此图谱的不同将它们区分开来。(3)对4例十二指肠溃疡伴Hp感染治疗前及治疗后1年再现的菌株的RAPD分析,发现3例患者的菌株具有相同的RAPD图谱,证实为原菌株的复发,而1例患者的菌株则产生不同的RAPD图谱,推测为新菌株的再感染,4例患者随访1年均为十二指肠溃疡复发。结论 RAPD分析只需单一的随机引物,方法简单、快速、经济、重复性好,可用于区分不同来源的Hp菌株,特别对治疗前后再现的菌株复发和再感染的鉴别更具临床应用价值。
【主题词】 螺杆菌， 幽门 多态现象(遗传学)DNA
Application of PCR based RAPD in differential diagnosis of strains of H.pylori Wang Weihong, Yan Zanhua, Hu Fulian. Department of Gastroenterology, Beijing Medical University First Teaching Hospital, Beijing 100034
【Abstract】 Objective To establish a rapid and sensitive polymerase chain reaction (PCR)based random amplified polymorphisc DNA (RAPD) fingerprinting method to distinguish different H.pylori isolates and evaluate its use in distinguishing recrudecence from reinfection of H.pylori after eradication. Methods H.pylori DNA was extracted and used for PCR based RAPD fingerprinting with a 10 nucleotide arbirary primer. The reproducibility and sensitivity of RAPD patterns was tested in 50 different clinical strains.Strains which isolated from 4 duodenal ulcer patients with H.pylori infection before treated with triple therapy and one year after eradication were tested with this method to confirm recrudecence or reinfection. Results RAPD fingerprinting yielded 10 to 20 main bands on the agarose gels.The profiles of the PCR products from the same isolates at two different times were indistinguishable. Fifty different clinical strains yielded different RAPD profile. By comparing 4 pairs of isolates before and after treatment,we confirmed that 3 patients with same RAPD profile belonged to recrudecence, and l patient with different RAPD profiles belonged to reinfection. Con[1] [2] [3] 下一页
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