CLONING OF THE PPU21IM GENE USING A IN-VIVO SELECTION METHOD

A genetic system enabling the in vivo selection of genes encoding the DNA-modifying enzymes was developed. A gene library is transformed int o a strain harboring the restriction-modification (R-M) system which a recognition sequence is a subset of the target sequence of the DNA me thyltransferase (MTase) to be cloned. If the residing MTase is tempera ture sensitive, the inability of transformants to grow at 42 degrees C provides a simple and convenient procedure for the isolation of new M Tase-encoding genes. The feasibility of this procedure has been demons trated by the isolation of the ppu21IM gene from a Pseudomonas putida RFL21 gene library.