Identifying stability of polymerase in master mixes used in PCR and repair possibilities for the degraded reagents

Abstract

The component of commercial available master mix most sensitive to unfavorable conditions is a polymerase. Available commercial polymerase chain reaction (PCR) master mixes are generally recommended for storage at −20°C, otherwise they may lose their activity. The aim of the experiment was to verify if storing mixes in adverse and extreme conditions may influence the quality of a PCR product. In the second phase of the research, it was to indicate if inactive PCR reagents that have lost their activity, may recover their enzymatic properties. Five different commercially available master mixes were incubated in unfavorable conditions. After the PCR, an electrophoresis was carried out and the obtained product was an evidence of a proper PCR reaction. Total degradation of mixes was caused by their incubation at room temperature for 28 days and incubation at 100°C for 60 minutes. Addition of polymerases to the degraded mixes (incubation at room temperature for 28 days) resulted in a regeneration of all of five mixes. In the case of polymerases incubated at 100°C for 60 minutes, regeneration was effective only in two of the five mixes. Our research confirms that PCR master mix is characterized by high resistance to varied conditions as well as in some cases can be repaired after degradation.