Zeng Fanwen, Zhong Wanhuan, Chen Tanzipeng, Wang Guoqian, Sa Jiaqi, Zhang Shouquan, Wei Hengxi, Chen Xuanjiao
Guangzhou Zoo & Guangzhou Wildlife Research Center Guangzhou China.
Kunming Institute of Zoology Chinese Academy of Sciences Kunming China.
Ecol Evol. 2024 Nov 19;14(11):e70551. doi: 10.1002/ece3.70551. eCollection 2024 Nov.
The difficulty in bird sex identification has made molecular sexing an important way to solve this problem. The conventional polymerase chain reaction (PCR) methods are time-consuming and dependent on laboratory equipment. Recombinase-aided amplification (RAA) is a rapid, specific, sensitive, and cost-effective isothermal nucleic acid amplification technique. Hence, a rapid birds sexing method based on real-time RAA targeting the unique conserved sequence 0.6-kb EcoRI fragment (EE0.6) gene of Carinatae birds has been established and showed good specificity at 39°C for 20 min. The limit of detection for the real-time RAA assay was determined to be 10 pg., which is 10 times more sensitive than the conventional PCR assay. For real clinical samples, the real-time RAA assay was successfully determined sex in a subset of nine bird species and was 100% consistent with the conventional PCR assay. Consequently, the present real-time RAA assay proves to be a powerful on-site detection tool that can be used for an efficient and reliable birds sexing for further studies on sex ratio and captive management.
鸟类性别鉴定的困难使得分子性别鉴定成为解决这一问题的重要方法。传统的聚合酶链反应（PCR）方法耗时且依赖实验室设备。重组酶辅助扩增（RAA）是一种快速、特异、灵敏且经济高效的等温核酸扩增技术。因此，基于实时RAA靶向鸡形目鸟类独特保守序列0.6-kb EcoRI片段（EE0.6）基因建立了一种快速的鸟类性别鉴定方法，该方法在39°C下20分钟显示出良好的特异性。实时RAA检测的检测限确定为10 pg，比传统PCR检测灵敏10倍。对于实际临床样本，实时RAA检测成功鉴定了9种鸟类中的部分样本的性别，且与传统PCR检测100%一致。因此，目前的实时RAA检测被证明是一种强大的现场检测工具，可用于高效、可靠的鸟类性别鉴定，以进一步研究性别比例和圈养管理。
