Thermally resistant nuclease in Serratia marcescens hinders PCR reactions and degrades PCR products

Polymerase chain reaction (PCR) is an important tool for exogenous gene acquisition and recombinants identification. There exist two problems when using Serratia marcescens as a template for PCR amplification: amplified PCR products are rapidly degraded, and the results of PCR amplification are unstable. The aim of the present work was to elucidate the reasons for this. By mixing PCR products amplified from Escherichia coli DH5α with S. marcescens supernatant or pellet, we found that the DNA‐degrading substance in S. marcescens is thermally resistant and present both intracellularly and extracellularly. We then determined that it is protein, and most likely S. marcescens nuclease, that degrades PCR products since the addition of SDS and EDTA can effectively inhibit or block the degradation of PCR products. By knocking out the S. marcescens nuclease encoding gene, nucA, we confirmed that the nuclease is responsible for the degradation of PCR products and the instability of PCR amplification. This work is the first to show that the S. marcescens nuclease is temporarily and partially inhibited by high temperatures during PCR and recovers rapidly at room temperature after PCR. Significance statement Amplified PCR products are rapidly degraded, and the results of PCR amplification are unstable when using Serratia marcescens as a template for PCR amplification. The present work confirmed that the nuclease is responsible for the degradation of PCR products and the instability of PCR amplification. It has been reported that S. marcescens nuclease is only active at temperatures from 0°C to 42°C, and there is no literature to date indicating the thermal stability of S. marcescens nuclease activity. There is even an article reporting that S. marcescens nuclease molecules are thermally degraded at temperatures above 40°C. This work is the first to show that the S. marcescens nuclease is thermally resistant.