Costantini V, Guaricci A C, Laricchiuta P, Rausa F, Lacalandra G M
Department of Animal Production, Faculty of Veterinary Medicine, University of Bari, Strada provinciale per Casamassima Km 3, 70010 Valenzano (BA), Italy.
Anim Reprod Sci. 2008 Jun;106(1-2):162-7. doi: 10.1016/j.anireprosci.2007.12.013. Epub 2008 Jan 3.
Humboldt Penguins (Spheniscus humboldti) show little sexual dimorphism, and although males are usually heavier and larger than females, sexing by direct observation may be difficult, especially in young subjects. In this paper we evaluate the utility of the molecular approach, for sexing impuberal Humboldt Penguins from feathers. Firstly, a PCR test was used employing primers that amplify the homologous region of the CHD-W gene, unique in female, and the CHD-Z gene, occurring in the two sexes. The analysis of the PCR products showed a band of 370 bp in males and two bands of 370 and 380 bp in females. Additionally, to confirm these results, the PCR products were digested with HaeIII and Asp700 for RFLP analysis. Male PCR products showed two bands (310 and 60 bp) after digestion with HaeIII, and a unique band (370 bp) using Asp700, while all fragments obtained from females resolved into three bands using both HaeIII (380, 310 and 60 bp) and Asp700 (370, 270 and 110 bp), confirming the previous PCR sex determination. Results from these two different DNA-based tests were in accordance, in all cases, with sexes checked by preliminary cloacoscopy. Thus, it was found that the PCR method from feather samples alone is sufficient, reliable and without any risks for a rapid sexing in Humboldt Penguin. This non-invasive sexing technique can be useful at any age to verify the sex ratio in field populations and for gender identification in ex situ conservation programs.
洪堡企鹅（Spheniscus humboldti）几乎没有明显的性别二态性，虽然雄性通常比雌性更重、体型更大，但通过直接观察来区分性别可能很困难，尤其是对于幼鸟。在本文中，我们评估了分子方法在从羽毛中鉴别未成年洪堡企鹅性别的实用性。首先，使用了一种PCR检测方法，该方法采用的引物能扩增CHD-W基因的同源区域（该基因仅存在于雌性中）以及CHD-Z基因（两性均有）。对PCR产物的分析显示，雄性有一条370 bp的条带，雌性有两条370和380 bp的条带。此外，为了证实这些结果，用HaeIII和Asp700对PCR产物进行消化以进行限制性片段长度多态性（RFLP）分析。雄性PCR产物用HaeIII消化后显示两条条带（310和60 bp），用Asp700消化后显示一条独特的条带（370 bp），而从雌性获得的所有片段用HaeIII（380、310和60 bp）和Asp700（370、270和110 bp）消化后均分解为三条条带，这证实了之前的PCR性别鉴定结果。在所有情况下，这两种基于DNA的不同检测结果均与通过初步泄殖腔镜检查确定的性别一致。因此，发现仅从羽毛样本进行PCR方法就足以快速、可靠地鉴别洪堡企鹅的性别，且没有任何风险。这种非侵入性的性别鉴定技术在任何年龄段都可用于核实野外种群的性别比例以及在迁地保护项目中进行性别鉴定。
