沉香随机扩增多态性DNA反应体系的建立  

Establishment of RAPD Reaction System of Aquilaria agallocha Roxb.

作　　者：邵敏[1] 周鹤峰[1] 唐历波[1] 刘银花[1] 李官成[1] 

机构地区：[1]遵义医学院珠海校区,广东珠海519041

出　　处：《时珍国医国药》2009年第1期74-76,共3页Lishizhen Medicine and Materia Medica Research

基　　金：遵义医学院青年科研基金(No.401203)

摘　　要：目的对影响沉香随机扩增多态性DNA（RAPD）反应的各因素进行优化，建立适用于沉香RAPD的最佳反应体系。方法采用改良的CTAB法提取基因组DNA，变化单一因素的方法对模板DNA浓度、Taq酶用量、dNTPs、引物浓度、Mg^2＋浓度、退火温度对RAPD影响进行分析。结果建立了适合沉香RAPD分析的最佳反应体系：在20μl反应体系中，模板DNA用量20ng、Taq酶1．0U、dNTPs浓度为0．3～0．4mmol·L^-1、随机引物浓度为0．6μmol·L、Mg^2＋浓度1．5—2．0mmol·L^-1、PCR反应条件：94℃，10min；94℃，1min；36℃，1min；72℃，2min；45个循环，72℃，8min。结论该反应体系具有良好的稳定性和重复性。Objective To establish the optimum RAPD reaction system of Aquilaria agaUocha Roxb. Methods The genomic DNA was extracted from Aquilaria agallocha Roxb. by modified CTAB method, and the different Template DNA, Taq, dNTPs, Primer, Mg^2＋, the annealing temperature were analyzed. Results The optimized reaction system and program of RAPD for Aquilaria agallocha Roxb. were as follows： The reactions were performed in a volume of 20μl,which containing 20ng DNA, 1U Taq ploymerase, 0.3～0.4 mmol· L^-1 dNTPs,0.6 μmol· L^-1 random Primer, 1.5 -2.0 mmol· L^-1Mg2＋. After pre -denaturing at 94℃ for 10 min, under the condition of denaturing at 94℃ amplifying for 45 cycles ,finally extension at 72℃ for 8 min. for 1 min,annealing at 36℃ for 1 rain,extension at 72℃ for 2 min, Conclusion The reaction system is reproducible and highly stable.

关 键 词：沉香 RAPD 基因组DNA 反应体系 

分 类 号：Q943.2[生物学—植物学] R2-03[医药卫生—中医学]