耐热FD DNA多聚酶在PCR中的应用条件

APPLICATION CONDITIONS OF THE THERMOSTABLE FD DNA POLYMERASE IN PCR

探索了用于多聚酶链式反应(PCR)的FD DNA多聚酶的最适反应条件在100μl的反应液中含有25 mmol/L的Tris-HCl(pH8.0),25 mmol/L的(NH_4)_2SO_4,1.5 mmol/L的MgCl_2,200 μg/ml的明胶,dNTP各200 μmol/L,模板量为3.3×10～(-2)Pg,引物量各为290 nmol/L,1～2 U的耐热FD DNA多聚酶,30次循环反应的系统为通用PCR最适反应系统。应用这一反应系统,混迹于大量DNA分子中的特定DNA片段可借助于琼脂糖凝胶电泳专一性地、快速而有效地检出。

Optimal conditions for PCR reaction using the FD DNA polymerase was inves-tigated. The optimal conditions were as follows: 25 mmol/L Tris-HCl (pH8.0), 25mmol/L(NH_4)_2SO_4. 1.5mmol/L MgCl_2, 200μg/ml Gelatin, 200μmol/L dNTPs, 3.3×10^(-2)pg template, 290 nmol/L for both primers and lor2 U thermostable FD DNApolymerase in 100μl reaction solution and 30 cycles of amplification were followedconventionally. With this reaction system minute amount of particular DNA frag-ment present among large amount of DNA could be detected specifically and effi-ciently with the aid of agarose electraphoresis.