THERMO SEQUENASE(TM) DNA-POLYMERASE AND T-ACIDOPHILUM PYROPHOSPHATASE- NEW THERMOSTABLE ENZYMES FOR DNA-SEQUENCING

A combination of thermostable enzymes has been developed that produces higher qualify Cycle sequences. Thermo Sequenase(TM) DNA polymerase i s a thermostable enzyme engineered to catalyze the incorporation of dd NTPs with an efficiency several thousandfold better than other thermos table DNA polymerases. Since the enzyme also catalyzes pyrophosphoroly sis at dideoxy termini, a thermostable inorganic pyrophosphatase is ne eded to remove the pyrophosphate produced during sequencing reactions. Thermoplasma acidophilum inorganic pyrophosphatase (TAP) is thermosta ble and effective for converting pyrophosphate to orthophosphate. The use of the combination of Thermo Sequenase polymerase and TAP for cycl e sequencing yields sequence data with uniform band intensities, allow ing the determination of longer; more accurate sequence reads. Uniform band intensifies also facilitate interpretation of sequence anomalies and the presence of mixed templates. Sequencing PCR products of DNA a mplified from heterozygous diploid individuals results in signals of e qual intensity from each allele.