Mycoplasma gallisepticum is the most important and infectious Mycoplasmosis. It is caused lots of economic losses for poultry's industry of Iran. The target of this study is comparison of culture and nested PCR techniques to detect Mycoplasma gallisepticum infection of chicken’s from industrial farms in Kerman province of Iran. 88 isolates received from industrial farms of Kerman province of Iran were measured by culture and nested PCR techniques. Two primers were used to identify Mycoplasma, but two pair’s primer for gallisepticum species. Mycoplasma galisepticum isolation and PCR results on samples from infected chickens received from Kerman province of Iran showed that PCR test was more sensetive than culture and this protocol can use as a remarkable way to diagnose Mycoplasma gallisepticum infection in birds. On the other hand, nested PCR is analytical test that its sensitivity showes two pairs of nested primer (the external primers and internal primers) can amplify two rigions of GapA gene and be more sensetive. Many routine tests DNA amplification methods have been developed for diagnosis of Mycoplasma gallisepticum and used in many in many laboratiries. PCR method based on the GapA can amplify DNA from Mycoplasma gallisepticum. The PCR method can that target surface protein gene. Nested gapA PCR is more sensetive test from PCR of GapA gene. This technique is a simple and fast method to detect and isolate infected birds, so it is a way to decrease economic losses in poultry industry. Four genes were recommended to Mycoplasma gallisepticum PCR that GapA gene is one of them that by nested PCR can detect Mycoplasma gallisepticum.