幽门螺杆菌cagA基因探针的制备与临床应用

Preparation of cagA gene probe and its clinical application

目的研究ｃａｇＡ＋Ｈｐ（幽门螺杆菌）与胃肠道疾病的关系，并探讨长臂光敏生物素标记的ｃａｇＡ基因探针的临床意义。方法构建含ｃａｇＡ基因片段的重组质粒克隆株，以重组质粒ＤＮＡ为模板ＰＣＲ扩增ｃａｇＡ基因片段用长臂光敏生物素标记产物ＤＮＡ，对病人进行基因探针检测及ＰＣＲ、尿酶试纸检测。结果浅表性胃炎、深度胃炎、胃溃疡、十二指肠溃疡、慢性萎缩性胃炎及胃癌病人的检测结果是：尿酶检测阳性百分率为：７０．３、７２．２、７７．７、６９．２、２５；ＰＣＲ阳性百分率分别为：５９．２、７７．８、７１．４、７０．７、７５、７８；基因探针杂交检测阳性率分别为：４８．１、６１．１、７０．３、７０．７、６６．７、６６．６、７２．２。结论ｃａｇＡ是胃、十二指肠疾病严重程度的标志，长臂光敏生物素标记的ｃａｇＡ基因探针具有特异性强、灵敏度高的特点。

Objective To investigate the relationship between cagA gene positive H.pylori and gastroduodenal diseases and the significance of the long arm photobiotin labelled cagA gene probe hybridizing detection in clinic. Methods We cloned the cagA gene fragment in pGEM T vector, and amplified cagA fragment with the template recombinant plasmid by PCR. Probes were prepared by labelling the fragment with the long arm photobiotin, and used to detect cagA + H.pylori in 137 patients, taking rapid urease test (RUT) and PCR as contrast. Results The positive percentage in superficial gastritis, deep gastritis, gastritic ulcer, duodenal ulcer, chronic atrophic gastritis and gastric cancer, (%) were 70 3, 72 2, 77 7, 69 2 and 25 respectively by URT, 59 2, 77 8, 71 4, 70 7, 75 and 78 respectively by PCR, and 48 1, 61 1, 70 3, 70 7, 66 7, 66 6 and 72 2 respectively by cagA gene probe detection. Conclusions Statistics suggest that cagA + H. pylori has a close correlation with gastroduodenal diseases in clinic, and can be a reliable predictor for high risk patients. The method of hybridizing detection with cagA gene probes, which labelled by the long arm photobiotin, is shown to be specific and sensitive , it is suitable for virulent strain typing identification in clinic.