Cao Xuan, Wang Ye-Fu, Zhang Chu-Fu, Gao Wen-Juan
College of Life Sciences, Wuhan University, Wuhan 430072, China.
Biosens Bioelectron. 2006 Sep 15;22(3):393-8. doi: 10.1016/j.bios.2006.05.011. Epub 2006 Jun 23.
Visual DNA microarrays, based on gold label silver stain (GLSS) and coupled with multiplex asymmetrical PCR, were developed for simultaneous, sensitive and specific detection of Ureaplasma urealyticum and Chlamydia trachomatis. 5'-end-amino-modified oligonucleotides, which were immobilized on glass surface, acted as capturing probes that were designed to bind complementary biotinylated targets DNA. The gold-conjugated streptavidins were introduced to the microarray for specific binding to biotin. The black image of microarray spots, resulting from the precipitation of silver onto nanogold particles bound to streptavidins, were used to detect biotinylated targets DNA visually or with a visible light scanner. Multiplex asymmetrical PCR of U. urealyticum, C. trachomatis and Bacillus subtilis (used as positive control) was performed to prepare abundant biotinylated single-stranded targets DNA, which affected detection efficiency and sensitivity of hybridization on microarray. Plenty of clinical samples of U. urealyticum and C. trachomatis from infected patients were tested using home-made DNA microarrays. For its high sensitivity, good specificity, simplicity, cheapness and speed, the present visual gene-detecting technique has potential applications in clinical fields.
基于金标银染（GLSS）并结合多重不对称PCR技术，开发了可视化DNA微阵列，用于同时、灵敏且特异检测解脲脲原体和沙眼衣原体。固定在玻璃表面的5'-端氨基修饰寡核苷酸作为捕获探针，其设计用于结合互补的生物素化靶标DNA。将金共轭链霉亲和素引入微阵列，使其与生物素特异性结合。银沉淀在与链霉亲和素结合的纳米金颗粒上，从而形成微阵列斑点的黑色图像，用于目视检测或用可见光扫描仪检测生物素化靶标DNA。对解脲脲原体、沙眼衣原体和枯草芽孢杆菌（用作阳性对照）进行多重不对称PCR，以制备大量生物素化单链靶标DNA，这会影响微阵列杂交的检测效率和灵敏度。使用自制的DNA微阵列对大量来自感染患者的解脲脲原体和沙眼衣原体临床样本进行检测。由于其高灵敏度、良好的特异性、简便性、低成本和快速性，这种可视化基因检测技术在临床领域具有潜在应用价值。
