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1 October 2009 A Simple and Improved DNA Test for Avian Sex Determination
Jae-Ik Han, Jeong-Ho Kim, Sukyung Kim, Shi-Ryoung Park, Ki-Jeong Na
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Determining the sex of a bird on the basis of its morphological characteristics is difficult in >50% of avian species; in nestlings, this rate is even higher. Given that male birds have two identical sex chromosomes (ZZ), whereas females are heterogametic (ZW), the development of a DNA-based sexing method has been widely investigated in several species. Griffiths et al. (1998) reported a simple DNA test using intronic size variation; however, this method cannot differentiate birds that have small or nonexistent size variations. We report a simple and novel DNA test for avian sex determination using a novel primer (p0) specific for the unique sequence of the chromohelicase-DNA-binding (CHD)-W gene. Use of a multiplex polymerase chain reaction (PCR) and simple agarose-gel electrophoresis with p0 and with the primers P2 and P8 described by Griffiths et al. (1998) revealed that the P0–P2 combination could amplify the partial CHD-W gene alone. Consequently, we found that p0 could be used to identify only the CHD-W gene. We anticipate that this multiplex PCR will be useful for universal avian sexing, regardless of the intronic size variation.

© 2009 by The American Ornithologists' Union. All rights reserved. Please direct all requests for permission to photocopy or reproduce article content through the University of California Press's Rights and Permissions website,,asp.
Jae-Ik Han, Jeong-Ho Kim, Sukyung Kim, Shi-Ryoung Park, and Ki-Jeong Na "A Simple and Improved DNA Test for Avian Sex Determination," The Auk 126(4), 779-783, (1 October 2009).
Received: 14 October 2008; Accepted: 20 April 2009; Published: 1 October 2009

CHD gene
CHD-W specific primer
sex differentiation
universal sexing
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