According to the invention, based on a pGA1 corynebacterium replicon in pEC-XK99E, which is a low-copy plasmid replicon with a copy number of about 30 in corynebacterium glutamicum, 325-site isoleucine of a replicating protein Rep is mutated into threonine, and 398-site amino acid for coding serine is changed into a synonymous codon, so that a replicon pGA1-Rep-I325T / S398 capable of increasing the plasmid copy number of corynebacterium is successfully obtained. The replicon is used for construction of high-copy plasmids, and after a plasmid pEC-XK99E-Rep-T / S carrying the pGA1-Rep-I325T / S398 replicon is replicated in corynebacterium glutamicum, it is proved that the number of plasmids in thalli is increased through plasmid in-vitro extraction. According to the method, pEC-XK99E-EGFP-Rep-T / S plasmids are constructed, fluorescence values under different thallus amounts are measured, and the fluorescence amount is increased by three times compared with that of mutant plasmids, namely, the expression amount of protein is increased by the modified plasmids. Meanwhile, by virtue of a fluorescent quantitative PCR means, the copy number after mutation is determined to be about 245 which is 8 times of that of a plasmid of a wild type replicon.