Enterococcus hirae is a Gram Positive Bacteria. Enterococcus hirae has an ability to produce Protease. In this experiment, we attempted for the protease producing gene in Enterococcus hirae and transferring the gene to non-Protease producing organism. Itmeanswhich gene that responsible to produce protease is identified by this experiment. Primer designing tools are used to design the specific primer for the amplification of DNA. The Primers are used for amplification. DNA is isolated from Enterococcus hirae using DNA isolationmethod. The isolated DNA is cross checked by the Agarose Gel Electrophoresis Method. The isolated DNA is further introduced into PCRmachine for amplification. The PCRMasterMix and Primers are used for amplification.After theDNAamplification, the cloning vectors are used for Cloning. The Ta plasmid vector (pBZ57RT) are used for DNA cloning, the cloned DNA with the vector is transformed into the Non protease producing organism, such as E.coli and checked for its activity.