SINGLE CHAIN BOVINE LUTEINIZING HORMONE AND METHOD FOR MANUFACTURING THE SAME

PURPOSE: Provided is a method for manufacturing single chain bovine luteinizing hormone(LH) which shows stronger hormone activity than one of natural LH, so as to be used for animal drugs. Thereby, LH is easily manufactured in a high yield as forming LH consisting of alpha and beta short fragments, into single chain LH. CONSTITUTION: A method for manufacturing single chain bovine luteinizing hormone(LH) comprises the following steps of: i) performing polymerase chain reaction to link DNA coding an alpha short fragment of bovine luteinizing hormone(LH) with DNA coding a beta short fragment of the hormone, using 4 kinds of oligonucleic acids bases as primers, to obtain cDNAs of the alpha and beta short fragments and cloning the cDNAs with pUC119; ii) multiplying cloned DNA through PCR to obtain bLH composed of cDNA of alpha and beta fragments, cutting the bLH by restriction enzyme Kpnl/Xbal and linking the bLH to vector pUC119 to form expression vector pcDAN3-bLH; iii) inserting the expression vector pcDAN3-bLH into CHO-K1 and cultivating CHO-K1 at media Ham F-12 including penicillin, streptomycin, glutamine, 10FCS and G418, in the presence of 5CO2 and air, at 37 deg.C for 2 weeks and selecting stable cells: iv) cultivating selected cells at 20 ml of CHO-S-SFM-11 media containing 50 units/ml of penicillin and 50 microg/ml of streptomycin at 37 deg.C for 48 hours, and centrifuging supernatant liquid obtained through the cultivation for 60 minutes to remove cell remnants.