Spargo C A, Haaland P D, Jurgensen S R, Shank D D, Walker G T
Becton Dickinson Research Center, Research Triangle Park, NC 27709.
Mol Cell Probes. 1993 Oct;7(5):395-404. doi: 10.1006/mcpr.1993.1058.
Strand displacement amplification, a new isothermal in vitro DNA amplification technique, was used to amplify target DNA contained within the IS6110 insertion element of the species within the Mycobacterium complex (Mycobacterium tuberculosis, M. bovis, M. bovis-BCG, M. africanum and M. microti). The target nucleic acid sequence is present in approximately ten, two, one, five and five copies in M. tuberculosis, M. bovis, M. bovis-BCG, M. africanum and M. microti, respectively. Amplified products were detected using a non-isotopic microtitre plate assay employing a biotinylated oligodeoxynucleotide probe and an alkaline phosphatase conjugated oligodeoxynucleotide probe. Lumiphos 530 was the chemiluminescent substrate for alkaline phosphatase. The combination of the strand displacement amplification method with this sensitive and rapid (less than 2 h) detection system resulted in the specific detection of as few as 1-25 initial IS6110 targets in the five Mycobacterium complex species based on signal/noise criteria. Negative results were obtained with eight other Mycobacterium species as well as with 32 non-Mycobacterium species.
链置换扩增是一种新的体外等温DNA扩增技术，用于扩增分枝杆菌复合群（结核分枝杆菌、牛分枝杆菌、卡介苗、非洲分枝杆菌和田鼠分枝杆菌）内各菌种IS6110插入元件中的靶DNA。靶核酸序列在结核分枝杆菌、牛分枝杆菌、卡介苗、非洲分枝杆菌和田鼠分枝杆菌中分别约有10、2、1、5和5个拷贝。使用生物素化寡脱氧核苷酸探针和碱性磷酸酶偶联寡脱氧核苷酸探针的非同位素微量滴定板测定法检测扩增产物。Lumiphos 530是碱性磷酸酶的化学发光底物。基于信号/噪声标准，链置换扩增方法与这种灵敏且快速（不到2小时）的检测系统相结合，能够特异性检测出五种分枝杆菌复合群菌种中低至1 - 25个初始IS6110靶标。对其他八种分枝杆菌菌种以及32种非分枝杆菌菌种检测结果均为阴性。
