To reveal the drug resistance and its mechanism preliminarily of clinical isolates of Corynebacterium striatum in Shandong Province of China, 83 isolates of Corynebacterium striatum were collected from clinical specimens during March 2017 and June 2020 in a tertiary hospitals from Shandong. The minimal broth dilution method was used to detect the minimal inhibitory concentration.The aminoglycoside modifying enzyme genes [aph(3”)-Ⅰb、aph(6)-Ⅰd、aac(6’)-Ⅰb], the ribosomal protective protein gene of tetracycline resistance (tetW), the ribosomal methylase gene (ermX), quinolone resistance genes (gyrA) were tested by polymerase chain reaction(PCR). The PCR products were sequenced and compared with BLAST.Quantitative real-time PCR (qPCR) was used to analyze the drug-resistant genes ermX and tetW quantitatively, and formula 2^－ΔΔCt was used to calculate the relative expression of genes.All the tested Corynebacterium striatum were proved as multi-drug resistant strains. The MDR-Cs showed completely sensitive to vancomycin and linezolid and completely resistance to erythromycin, ciprofloxacin and clindamycin, the detection rate of tetracycline, gentamicin were 86.7% (72/83)and 38.6%(32/83), respectively. The detection rate of ermX and tetW were 100%(83/83) and 88%(73/83), the detection rate of aph(3”)-Ⅰb, aph(6)-Ⅰd, aac(6’)-Ⅰb were 36.1%(30/83), 37.3%(31/83) and 15.7%(13/83), respectively. The resistance phenotype of the strain to erythromycin is cMLS. The drug resistance spectrum is relatively broad in this study, which needs to be paid attention from hospitals and laboratories.