栉孔扇贝RAPD反应体系的优化

Optimization of Condition for RAPD in Chlamys farreri

在研究栉孔扇贝不同地理野生种群的遗传多样性时 ,对RAPD反应中的模板浓度、引物浓度、dNTPs浓度、镁离子浓度等影响反应结果的因素进行了系统分析 ,并在此基础上 ,建立了栉孔扇贝RAPD实验室研究的最佳反应体系 :2 5 μl反应体系中 ,模板浓度为 2 5ng ,引物浓度为 2 5ng,dNTPs为 10 0 μM ,TaqDNA酶 1个单位 ,10×Buffer 2 5 μl,镁离子浓度 2mM .PCR循环参数为 :94℃预变性 5min ,然后 94℃变性 0 5min ,36℃退火 1min ,72℃延伸 1min ,共进行 45个循环 ,最后 72℃延伸 10min .

The conditions of RAPD for genetic diversity of different geography wildness population in Chlamys farreri were optioned.For a 25 μl reaction,the optional composition included 25 ng of template DNA,25 ng of random primers,100 μM dNTPs,1 U Taq DNA,10×Buffer 2.5 μl and 2 mM MgCl.The optional program for efficient amplification included 45 cycles of denaturation at 0.5 min(94 ℃ for 5 min for the first cycle),annealing at 36 ℃ for 1 min and extension at 72 ℃ for 2 min(10 min for the final cycle).