DNA PROBES FOR THE DIAGNOSIS OF PERIODONTAL PATHOGENS

特殊细菌在牙周炎和牙周炎中的作用 牙周炎得到了越来越多的科学文献的支持。然而， 传统的微生物技术限制了疾病研究的进展 诊断和监测。这个第一阶段项目的目的是 从与细菌相关的染色体DNA中开发DNA探针 牙周炎和牙周炎。探针将被标记为32P和 与固定在硝基纤维素上的细菌DNA杂交。这个 每个探针的杂交特异度将在标准中进行评估 氯化钠溶液和一种新的盐溶液--四乙基氯化铵 (TEACl)。这一新的盐制取消了A-T的优先熔化 相对于DNA杂交体中的G-C碱基对，因此产生了显著的 熔融轮廓的锐化(在TEACl中低于1C，而在氯化钠中为5-10C) 提高了探针的特异性。TEACl盐中的DNA探针 系统将允许开发高度敏感和特定的DNA 病原菌的检测、计数和监测的探针分析 细菌。该项目的长期目标是提供一个简单、 快速、准确、低成本的检测和检测方法 患者临床分离株细菌的鉴定 患有牙周炎或牙周炎。对特定病原体的诊断试验 将被开发并商业化生产，用于临床诊断 实验室和牙科诊所。

The role of specific bacteria as the etiologic agents of gingivitis and periodontitis is supported by a growing scientific literature. However, conventional microbiological techniques have restricted advances in disease diagnosis and monitoring. The purpose of this Phase I project is to develop DNA probes from the chromosomal DNA of bacteria associated with gingivitis and periodontitis. The probes will be labeled with 32P and hybridized against bacterial DNA immobilized on nitrocellulose. The hybridization specificity of each probe will be evaluated in the standard NaCl solution, and in a new salt solution, tetraethylammonium chloride (TEACl). This new salt system abolishes the preferential melting of A-T versus G-C base pairs in DNA hybrids, thus producing a significant sharpening of the melting profile (less than 1C in TEACl vs. 5-10C in NaCl) and enhancing the specificity of the probes. DNA probes in a TEACl salt system will allow the development of highly sensitive and specific DNA probe assays for the detection, enumeration and monitoring of pathogenic bacteria. The long-term objective of this project is to provide a simple, rapid, accurate and low-cost methodology for the detection and identification of the bacteria present in clinical isolates from patients with gingivitis or periodontitis. Diagnostic tests for specific pathogens will be developed and produced commercially for use by clinical diagnostic laboratories and in the dental office.