Background: Sex determination in birds is essential for effective breeding programs, conservation efforts, and scientific studies. However, the lack of visible sexual dimorphism in many species presents challenges, especially for nestlings. To address this issue, the most reliable methods of sex determination utilize the unique properties of the Z and W sex chromosomes. These features can be identified through polymerase chain reaction (PCR) techniques, allowing to differentiate between males (ZZ) and females (ZW). This study aimed to evaluate the effectiveness of PCR-based sex determination in a range of bird species. Methods: A total of ۲۷۰ blood and feather samples were collected from diverse bird species from Tehran province. Samples were collected from various parrot species, such as cockatiel (Nymphicus hollandicus), african grey parrot (Psittacus erithacus), and alexandrine parakeet (Psittacula eupatria), as well as from owls (Strigiformes), cranes (Gruiformes), common myna (Acridotheres tristis), and diamond dove (Geopelia cuneata).
PCR analysis was conducted using the primer pair ۲۵۵۰F and ۲۷۱۸R, designed to amplify the
CHD gene variants located on the Z and W sex chromosomes. The CHD۱Z and CHD۱W bands were analyzed to differentiate males (ZZ) from females (ZW). Results: Out of ۲۷۰ samples, ۱۰۳ males and ۱۴۱ females were successfully identified; nevertheless, there were challenges in accurately identifying ۲۶ birds. The selected primer pair successfully identified the sex of the majority. However, it was found ineffective in identifying the sex of Psittacula eupatria, Geopelia cuneate, and Psittacula krameri indicating a limitation in its applicability for this particular group. Conclusion: Given these findings, it is recommended that alternative primer pairs be explored for accurate sex determination in alexandrine parakeets, rose-ringed parakeets, and diamond doves. The results highlight the importance of using species-specific approaches in molecular sexing methods to enhance accuracy and reliability.