随机引物PCR检测人DNA指纹条件的优化  

Study on Optimal Conditions in Arbitrarily Primed PCR Human DNA Fingerprinting

作　　者：童大跃[1] 徐平[1] 区敬华[2] 郭语彬[1] 李芳[1] 林达[1] 伍新尧[2] 

机构地区：[1]中山医科大学生物化学教研室,广东广州510089 [2]中山医科大学法医学系,广东广州510089

出　　处：《中山医科大学学报》2001年第3期231-234,共4页Academic Journal of Sun Yat-sen University of Medical Sciences

基　　金：国家自然科学基金资助项目 ( 39370 75 5 );广东省重点科技攻关项目 ( 1997);广州市科委资助项目 (JB0 2 97 Z 44 0 2 )

摘　　要：【目的】探讨随机引物PCR检测人DNA指纹技术的最优化条件。【方法】应用随机引物PCR技术 ,用一对经筛选的短引物 ,对扩增人DNA指纹的条件进行了优化。【结果】实验结果显示 :DNA模板应新鲜 ,质量浓度在 5 0～ 5 5 0mg/L均有扩增产物。dNTP的浓度 0 2mmol/L最宜。Mg2 + 浓度 5 0mmol/L效果最佳。循环参数以两个三步PCR ,变性温度分别用 94℃、90℃ ,时间各为 30s,退火温度用 43℃、48℃ ,时间分别为 40s、5 0s,延伸温度用 72℃ ,时间分别为 1min、1min 2 0s。【结论】按照优化的实验条件 ,得出的DNA指纹图重复性好 ,个体特异性高 。To explore the optimal conditions in fingerprinting (APHDF). The human DNA fingerprints were detected by APHDF.A pair of short primers was used for amplification. The experimental conditions including template, Mg 2+ , deoxyribonucleotides, and parameters of cycle, were optimized. The template DNA should be abstracted freshly and the concentration should be ranged from 50～550 mg/L.The best concentration of Mg 2+ was 5 0 mmol/L. The deoxyribonucleotides concentration was optimal at 0 2 mmol/L. The PCR cycling parameters were as follows :The denaturing temperatures,annealing temperatures and extension temperatures were 94 ℃ and 90 ℃ for 30 s, 43 ℃ and 48 ℃ for 40 s or 50 s,and 72 ℃ for 1 min or 80 s, respectively.[Conclusion] The optimal conditions of the experiment are obtained, with good reproducibility and high specificity. Therefore, this method can be widely applied in practice.

关 键 词：AP-PCR DNA指纹法 聚合酶链反应 随机引物 

分 类 号：Q523.03[生物学—生物化学]