[利用尼龙膜通过DNA探针特异性分离大肠杆菌O157并检测其与志贺毒素大肠杆菌（STEC）组的关系]
[Utilization of nylon membranes for the specific isolation of Escherichia coli O157 with DNA probes and checking for relationship to the shiga toxin Escherichia coli (STEC) group].
作者信息
Gallien P, Much C, Perlberg K W, Protz D
机构信息
Bundesinstitut für gesundheitlichen Verbraucherschutz und Veterinärmedizin in Dessau.
出版信息
Berl Munch Tierarztl Wochenschr. 1999 Feb;112(2):58-63.
A method for specific isolation of Escherichia coli strains of serotype O157 is given. DNA-hybridization technique by using DIG-labeled specific O157 PCR-amplificates as probes is the basis. These investigations can be used for the detection and isolation of shigatoxin-producing Escherichia coli (STEC) of this serotype in combination with other tests for detection of shigatoxin genes (stx). No background is seen by using 'DIG Easy Hyb' solution and Nylon membranes for colony-and plaque-hybridization (Roche Diagnostics Boehringer Mannheim). Dark brown spots (E. coli colonies) are visible on the membranes after staining. After comparing the membrane with the masterplate it is possible to isolate the colonies from the masterplate, respectively. It is necessary to make proof that the isolates possess the rfb-genes (verification of serotype O157) and the stx-genes (test for belonging to the STEC-group). Both tests could be done by using PCR. In case of positive results other virulence factors of the isolates can be detected.
给出了一种特异性分离O157血清型大肠杆菌菌株的方法。以使用地高辛（DIG）标记的特异性O157 PCR扩增产物作为探针的DNA杂交技术为基础。这些研究可与其他检测志贺毒素基因（stx）的试验相结合，用于检测和分离该血清型产志贺毒素大肠杆菌（STEC）。使用“DIG Easy Hyb”溶液和尼龙膜进行菌落和噬菌斑杂交（罗氏诊断宝灵曼）时未见背景干扰。染色后，膜上可见深棕色斑点（大肠杆菌菌落）。将膜与原平板进行比较后，即可分别从原平板上分离出菌落。有必要证明分离株具有rfb基因（O157血清型验证）和stx基因（检测是否属于STEC组）。这两项检测均可通过PCR进行。如果结果为阳性，则可检测分离株的其他毒力因子。
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