Polymerase Chain Reaction (PCR) using MG 14F and MG 13R specific primers for Mycoplasma gallisepticum (MG) according to the promulgation of OIE (OIE., 2008) has successfully been applied for detecting chronic respiratory disease (CRD) caused by MG in domestic chickens. The diagnostic results for CRD by using PCR technique indicated that 20/43 (46.5 percent) of trachea and lung tissue samples of chickens suspecting Mycoplasma gallisepticum infection were positive. The size of specific PCR product was 185 bp. In this study, the lung tissues samples of CRD suspected chickens were also used for isolation of MG in Mycoplasma Agar (MA) and Mycoplasma Broth (MB) medium. The studied results showed that 89/174 (51.15 percent) samples were positive for M B media while 72/174 (41.38 percent) samples were positive for MA media.