Cloning of DNA Polymerase I Geobacillus thermoleovorans SGAir0734 from a Batu Kuwung Hot Spring in Escherichia coli

Access to biological engineering has become a critical point of modern science development through polymerase chain reaction (PCR). One of the main components in this process is DNA polymerase, which copies the main template DNA. However, there is a lack of studies on the production of DNA polymerase from indigenous thermophilic bacteria in Indonesia. To examine this process, DNA polymerase I gene (DNA pol I) from Geobacillus thermoleovorans (isolated from Batu Kuwung, Banten, Indonesia) was transformed into Escherichia coli. The gene was cloned by the cut and ligation method using NcoI and BamHI restriction enzymes, which were ligated with a pET23d vector. The recombinant gene was overexpressed in E. coli BL21 and identified by using SDS-PAGE of 10% acrylamide gel, showing that the protein molecular weight was approximately 100 kDa. This study successfully amplified the gene of interest, indicated by a high local similarity between the sequencing results and theoretical gene and positive intercellular protein expression. The results confirm that the study successfully cloned and synthesized recombinant DNA pol I of Geobacillus thermoleovorans from Batu Kuwung, Serang, Banten.