Mg^(2+)浓度在多基因PCR中关系的研究

Relationship between magnesium and multiplex polymerase chain reaction

目的 通过实验介绍游离性Mg2 + 在不同浓度情况下对Tag酶活性及MgCl2 作为一种盐对多基因PCR的影响过程。方法 利用不同Mg2 + 浓度的缓冲液及不含KCl的不同Mg2 + 浓度的缓冲液来研究其对多基因PCR的影响。结果 Mg2 + 是Tag酶活性所必需的 ,Mg2 + 浓度过低时 ,Tag酶活性显著下降 ,无特异条带扩增或特异条带扩增不明显。在Mg2 + 浓度高时 ,Tag酶活性增强 ,非特异条带扩增。随着Mg2 + 浓度的增加 ,在接近 10mmol/L时 ,扩增条带亮度减弱 ,非特异扩增条带减少。结论 在无KCl存在 ,单一Mg2 + 影响反应的情况下 ,随着Mg2 + 浓度的过度增加 ,高浓度的Mg2 + 抑制了Tag酶活性 ,从而减少了产物的数量 ,在 5 0mmol/LMg2 + 浓度时完全抑制了扩增。

Objective To get the relationship between magnesium and multiplex polymerase chain reaction.Methods With different magnesium concentration of buffer and with different magnesium concentration of buffer that without KCl to perform multiplex polymerase cthain reaction.Results At the higher magnesium concentration unspecific products appeared,but they gradually decreased in intensity towards 10 mmol/L magnesium concentrations.At 5 mmol/L magnesium concentration without KCl salt,there were specific products,but increasing with the magnesium concentration,at 50 mmol/L,the multiplex polymerase chain reaction was completely inhibited,there were no specific products.Conclusion The concentration of magnesium was necessary for Tag DNA polymerase.Tag DNA polymerase required free magnesium.2 mmol/L concentration of magnesium was very best for the five genes multiplex polymerase chain reaction.