RAPID SIMPLIFIED PROTOCOL FOR PURIFICATION OF TAQ DNA POLYMERASE FRAGMENT EXPRESSED IN ESCHERICHIA COLI.

An economical and simple method, boiling lysis method was developed and used to purify 94kDa Taq DNA polymerase fragment from transformed BL21 (DE3) cells. It is based on conditions such as thermostable properties, boiling time, removal of nucleic acids to achieve a high yield and activity of Taq DNA polymerase fragment on one cycle of boiling for 60mins. The centrifugation protocol followed was ideal for the purification, followed by streptomycin sulphate treatment to remove nucleic acids. The clear supernatant containing heat resistant Taq DNA polymerase was separated and stored at -70°C. The activity of enzyme was compared with commercial Taq DNA polymerase, stored in buffer containing 50% glycerol, at -20°C. The purified enzyme has a molecular weight of 94kDa, as predicted by SDS-PAGE and yielded appropriate enzyme activity comparing to the commercial Taq DNA polymerase. The whole process of purification was achieved within 3hrs. A total 53mg of Taq DNA polymerase enzyme was purified from 1 liter of bacterial culture emphasizing the fact that our method was economical and simple.