随机引物PCR检测人DNA指纹条件的优化

【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50～550mg/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、50s，延伸温度用72℃，时间分别为1min、1min20s。【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，为APHDF的推广提供了实验基础。

Study on Optimal Conditions in Arbitrarily Primed PCR Human DNA Fingerprinting

【Objective】To explore the optimal conditions in fingerprinting (APHDF).【Methods】The human DNA fingerprints were detected by APHDF.A pair of short primers was used for amplification.The experimental conditions including template,Mg2+,deoxyribonucleotides,and parameters of cycle,were optimized.【Results】The template DNA should be abstracted freshly and the concentration should be ranged from 50～550 mg/L.The best concentration of Mg2+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.