摘要
目的探索一种快速、有效且对人体无创伤的基因组DNA提取方法。方法比较碘化钾法和口腔拭子基因组DNA提取试剂盒法对新鲜和室温放置1周的唾液标本的基因组DNA提取和TP53、PRB-3基因PCR扩增效果。同时采集99名健康儿童和成人的唾液标本验证碘化钾法提取唾液基因组DNA的稳定性。结果两种DNA提取方法都能从新鲜唾液标本中获得高质量的基因组DNA,其中碘化钾法的得率和D260/D280分别为(1.91±0.15)μg和1.99±0.05,试剂盒法分别为(2.64±0.34)μg和1.81±0.02。对于室温放置1周的唾液标本,两种提取方法获得的DNA虽然降解明显,但是对TP53和PRB-3基因都能扩增正确大小的目的片段。碘化钾法提取的99名健康儿童和成人的唾液基因组DNA虽然个体差异明显[得率为(1.89±0.46)μg],但是DNA质量稳定可靠(D260/D280为1.96±0.10)。结论碘化钾法提取唾液基因组DNA不仅廉价、高效,而且对人体无创伤,非常适合大范围分子流行病学研究。
Objective To explore a fast, effective and noninvasive method of genomic DNA extraction. Methods We compared the genomic DNAs extracted by two DNA extraction methods of potassium iodide （KI） and Kit from saliva （fresh and placed a week at room temperature）, and PCR amplifications were done for TP53 and PRB-3 genes from their extracted DNA. Ninety-nine healthy children and adults were recruited and their saliva samples were collected to detect the stability of DNA extracted by KI. Results High quality genomic DNAs were extracted by both methods from the fresh saliva. The DNA yield based on KI was （1.91±0. 15）μg, with the D26o/D28o being 1. 99±0. 05, and that of Kit was （2. 64±0. 34）μg, with the D26o/ D28o being 1.81±0. 02. Although the extracted DNAs had obvious degradation from saliva samples placed for a week at room temperature, the expectant DNA fragments were successfully amplified by PCR for TP53 and PRB-3 genes using these extracted DNAs. Although the genomic DNAs from saliva samples of 99 healthy volunteers by the method of KI showed individual difference （DNA yield was El. 89±0.46]μg）, the DNA quality was stable and reliable （D260/D2so was 1.96±0. 10）. Conelusion KI method for extracting genomic DNA from saliva is not only cheap and highly effective, but also noninvasive, making it suitable for large-scale epidemiological studies.
出处
《第二军医大学学报》
CAS
CSCD
北大核心
2015年第5期548-553,共6页
Academic Journal of Second Military Medical University
基金
国家自然科学基金(81102703)
广东省科技计划项目(2013A032500005)
广东省中医药管理局项目(20123001)
广东药学院基础学院教改项目~~
关键词
唾液
碘化钾DNA提取
聚合酶链反应
saliva
potassium iodide
DNA extraction
polymerase chain reaction
