A recombinant vector comprising an incorporated chromosome DNA fragment containing a PvuI restriction enconuclease gene derived from Proteus vulgaris, a host transformed with the recombinant vector and a method of producing PvuI restriction endonuclease characterized in that the transformed host is cultivated and PvuI restriction endonuclease is harvested from the resulting culture. Since the host transformed with the recombinant vector of the present invention produces PvuI alone, it is unnecessary to remove PvuII in the purification process for PvuI, and further, since its productivity for nonspecific DNase is lower in comparison with conventional producer bacteria, DNase can be removed easily, making the production of PvuI easy.