Visualization of Sex Identification in Red‐Crowned Crane (Grus japonensis) via Recombinase‐Aided Amplification Combined With Pyrococcus furiosus Argonaute Assay
ABSTRACT The red‐crowned crane (Grus japonensis), a Class I protected animal in China, inhabits Northeast Asia, including China, Russia, and Japan. As sex‐monomorphic birds, red‐crowned cranes cannot be directly distinguished between females and males through observation. Molecular methods are accur...
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Wiley
2025-07-01
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| Series: | Ecology and Evolution |
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| Online Access: | https://doi.org/10.1002/ece3.71780 |
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| author | Shenluan Tan Tongtong Zhan Fanwen Zeng Xuanjiao Chen Tanzipeng Chen Li Li Hengxi Wei Shouquan Zhang Kejing Zuo |
| author_facet | Shenluan Tan Tongtong Zhan Fanwen Zeng Xuanjiao Chen Tanzipeng Chen Li Li Hengxi Wei Shouquan Zhang Kejing Zuo |
| author_sort | Shenluan Tan |
| collection | DOAJ |
| description | ABSTRACT The red‐crowned crane (Grus japonensis), a Class I protected animal in China, inhabits Northeast Asia, including China, Russia, and Japan. As sex‐monomorphic birds, red‐crowned cranes cannot be directly distinguished between females and males through observation. Molecular methods are accurate and stable for sex identification in birds and are widely used in zoos and farms. With the development of isothermal techniques, recombinase‐aided amplification (RAA) has provided novel insights into bird sexing owing to its low equipment dependence and rapid amplification. Advancements in the Pyrococcus furiosus Argonaute (PfAgo) biosensor have facilitated clinical detection. In this study, an innovative sex identification system was developed by integrating RAA and PfAgo in red‐crowned cranes. The RAA‐PfAgo system identified both females and males with remarkable accuracy. Via proper design of primers set, gDNA and probe, the RAA‐PfAgo system can complete visual detection, with detection limits between 0.35 ng/μL and 0.035 ng/μL under optimal conditions. The test samples exhibited strong green fluorescence in females, whereas no fluorescence was observed in males under blue light. The results of RAA‐PfAgo in the field were consistent with those obtained using conventional PCR. This study provides a high degree of rapidity, accuracy, and sensitivity for the sex identification of red‐crowned cranes. |
| format | Article |
| id | doaj-art-3a1d924679974f49affe0a8bd90946e6 |
| institution | Kabale University |
| issn | 2045-7758 |
| language | English |
| publishDate | 2025-07-01 |
| publisher | Wiley |
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| series | Ecology and Evolution |
| spelling | doaj-art-3a1d924679974f49affe0a8bd90946e62025-08-20T03:58:44ZengWileyEcology and Evolution2045-77582025-07-01157n/an/a10.1002/ece3.71780Visualization of Sex Identification in Red‐Crowned Crane (Grus japonensis) via Recombinase‐Aided Amplification Combined With Pyrococcus furiosus Argonaute AssayShenluan Tan0Tongtong Zhan1Fanwen Zeng2Xuanjiao Chen3Tanzipeng Chen4Li Li5Hengxi Wei6Shouquan Zhang7Kejing Zuo8State Key Laboratory of Swine and Poultry Breeding Industry, National Engineering Research Center for Breeding Swine Industry, Guangdong Provincial Key Lab of Agro‐Animal Genomics and Molecular Breeding, College of Animal Science South China Agricultural University Guangzhou ChinaBeijing Zoo, Beijing Key Laboratory of Captive Wildlife Technologies Beijing ChinaGuangzhou Zoo & Guangzhou Wildlife Research Center Guangzhou ChinaGuangzhou Zoo & Guangzhou Wildlife Research Center Guangzhou ChinaGuangzhou Zoo & Guangzhou Wildlife Research Center Guangzhou ChinaState Key Laboratory of Swine and Poultry Breeding Industry, National Engineering Research Center for Breeding Swine Industry, Guangdong Provincial Key Lab of Agro‐Animal Genomics and Molecular Breeding, College of Animal Science South China Agricultural University Guangzhou ChinaState Key Laboratory of Swine and Poultry Breeding Industry, National Engineering Research Center for Breeding Swine Industry, Guangdong Provincial Key Lab of Agro‐Animal Genomics and Molecular Breeding, College of Animal Science South China Agricultural University Guangzhou ChinaState Key Laboratory of Swine and Poultry Breeding Industry, National Engineering Research Center for Breeding Swine Industry, Guangdong Provincial Key Lab of Agro‐Animal Genomics and Molecular Breeding, College of Animal Science South China Agricultural University Guangzhou ChinaGuangzhou Zoo & Guangzhou Wildlife Research Center Guangzhou ChinaABSTRACT The red‐crowned crane (Grus japonensis), a Class I protected animal in China, inhabits Northeast Asia, including China, Russia, and Japan. As sex‐monomorphic birds, red‐crowned cranes cannot be directly distinguished between females and males through observation. Molecular methods are accurate and stable for sex identification in birds and are widely used in zoos and farms. With the development of isothermal techniques, recombinase‐aided amplification (RAA) has provided novel insights into bird sexing owing to its low equipment dependence and rapid amplification. Advancements in the Pyrococcus furiosus Argonaute (PfAgo) biosensor have facilitated clinical detection. In this study, an innovative sex identification system was developed by integrating RAA and PfAgo in red‐crowned cranes. The RAA‐PfAgo system identified both females and males with remarkable accuracy. Via proper design of primers set, gDNA and probe, the RAA‐PfAgo system can complete visual detection, with detection limits between 0.35 ng/μL and 0.035 ng/μL under optimal conditions. The test samples exhibited strong green fluorescence in females, whereas no fluorescence was observed in males under blue light. The results of RAA‐PfAgo in the field were consistent with those obtained using conventional PCR. This study provides a high degree of rapidity, accuracy, and sensitivity for the sex identification of red‐crowned cranes.https://doi.org/10.1002/ece3.71780recombinase‐aided amplification‐Pyrococcus furiosus Argonaute (RAA‐PfAgo)red‐crowned cranesex identification |
| spellingShingle | Shenluan Tan Tongtong Zhan Fanwen Zeng Xuanjiao Chen Tanzipeng Chen Li Li Hengxi Wei Shouquan Zhang Kejing Zuo Visualization of Sex Identification in Red‐Crowned Crane (Grus japonensis) via Recombinase‐Aided Amplification Combined With Pyrococcus furiosus Argonaute Assay Ecology and Evolution recombinase‐aided amplification‐Pyrococcus furiosus Argonaute (RAA‐PfAgo) red‐crowned crane sex identification |
| title | Visualization of Sex Identification in Red‐Crowned Crane (Grus japonensis) via Recombinase‐Aided Amplification Combined With Pyrococcus furiosus Argonaute Assay |
| title_full | Visualization of Sex Identification in Red‐Crowned Crane (Grus japonensis) via Recombinase‐Aided Amplification Combined With Pyrococcus furiosus Argonaute Assay |
| title_fullStr | Visualization of Sex Identification in Red‐Crowned Crane (Grus japonensis) via Recombinase‐Aided Amplification Combined With Pyrococcus furiosus Argonaute Assay |
| title_full_unstemmed | Visualization of Sex Identification in Red‐Crowned Crane (Grus japonensis) via Recombinase‐Aided Amplification Combined With Pyrococcus furiosus Argonaute Assay |
| title_short | Visualization of Sex Identification in Red‐Crowned Crane (Grus japonensis) via Recombinase‐Aided Amplification Combined With Pyrococcus furiosus Argonaute Assay |
| title_sort | visualization of sex identification in red crowned crane grus japonensis via recombinase aided amplification combined with pyrococcus furiosus argonaute assay |
| topic | recombinase‐aided amplification‐Pyrococcus furiosus Argonaute (RAA‐PfAgo) red‐crowned crane sex identification |
| url | https://doi.org/10.1002/ece3.71780 |
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