Preparation And Application Of Recombinant Nuclease

Extracellular nuclease from Serratia marcescens is a saccharide nonspecific endonucleases which can degrade DNA and RNA.whether it is single-stranded, double-stranded,linear,cyclic,or supercoiled form. The nuclease has important application value in the purification of proteins and other biological products,in electrophoresis and chromatography sample processing and in removing nucleic acid residues from recombinant biological products.Gene of S.marcescens nuclease(nsnu) was prepared by PCR with genome extracted from S.marcescens. Expression vector pET-22b-nsnu was constructed by the ligation of plasmid pET-22b with the gene of nsnu and sequenced.Recombinant nuclease was expressed in E.coli induced by IPTGThe best expression condition was settled. Recombinant nuclease was purified by His-Binding-resin which resulted in10mg Soluble protein per1L culture medium.and its Purity was above95%.We detected The activity of purified recombinant nuclease which was1.04×106U/mg.The activity of purified recombinant nuclease in different conditions was tested.The results showed that purified recombinant nuclease could maintain its activity in a large range of conditions.We applied the recombinant nuclease in production of L-Methionine y-lyase in our lab.The application results showed that very small amounts of recombinant nuclease can greatly reduce the fracture fluid viscosity,which eliminated the ultrasonic treatment process and shortened the separation time.The use of recombinant nuclease could apparently separate precipitation and supernatant in which way simplified purification steps.This work indicates the recombinant nuclease has widely potential efficacy in future application.