Sex identification of Crested Serpent Eagle and Collared Scops Owl using random amplified polymorphic DNA fingerprinting

• Main Authors: Hsin-An Hsu, 許欣安
• Other Authors: Shih-Torng Ding
• Format: Others
• Language: zh-TW
• Published: 2008
• Online Access: http://ndltd.ncl.edu.tw/handle/41603977179501230041

碩士 === 國立臺灣大學 === 動物科學技術學研究所 === 96 === Crested Serpent Eagle (Spilornis cheela hoya), Collared Scops Owl (Otus bakkamoena), Crested Goshawk (Accipiter trivirgatus) Tawny Fish Owl (Ketupa flavipes) and Grass Owl (Tyto longimembris) are sexually monomorphic birds. To identify their sexes by their apparent morphology is difficult. Such characteristic complicates the research on establishing breeding program and conservation for these animals. Therefore, the objective of this study was to investigate sex-specific genetic markers of Crested Serpent Eagle and Collared Scops Owl using random amplified polymorphic DNA (RAPD) fingerprinting. we used sex-specific primers of avian CHD1 (chromo-helicase-DNA-binding 1) gene and EE0.6 (EcoRI 0.6-kb fragment) sequence to identify these five birds sex. Besides, 120 random primer sequences were from RAPD 10 mers kits (Operon Biotechnologies) and used for RAPD fingerprinting to search for sex-specific fragments of Crested Serpent Eagle and Collared Scops Owl. The results showed that CHD1 gene could correctly identify sex in Crested Serpent Eagle and Grass Owl only. Furthermore, EE0.6 sequence could also correctly distinguish sex of these birds, the useful primers sets are A, C and D for Crested Goshawk; E. primers sets for Collared Scops Owl and Tawny Fish Owl; C、D and F for Grass Owl. Moreover, The RAPD results showed that OPBB08 and OPAT15 random primers generated approximately 1.2 and 1.3 kb sex-specific fragments in female Crested Serpent Eagle and OPAN15 random primer generated a 1.3 kb sex-specific fragment in female Collared Scops Owl. The sex-specific fragments were constructed into the vectors for sequencing. According to the sex-specific fragments sequences, several primers were designed for sex identification in 41 Crested Serpent Eagle and 16 Collared Scops Owl respectively. The CseBB08-5a (Crested serpent eagle BB08-5a) and CseBB08-5b primers amplified 553 and 895 bp fragment respectivily in all female Crested Serpent Eagle but not in the male; the CseBB08-7 primers amplified a 194 bp fragment in all female Crested Serpent Eagles and a 543 bp fragment in all females and males; the CseAT15 primers amplified a 734 bp fragment in all female Crested Serpent Eagle and a 147 bp fragment in all females and males. The CsoAN15-6 (Collared scops owl AN15-6) primers from Collared Scops Owl genomic sequences amplified a 349 bp fragment in all female Collared Scops Owl and a 796 bp fragment in all females and males. The common amplified fragment in both male and female birds can be used as an internal control. In addition, we tested those primers on other species and found that CseBB08-5b of Crested Serpent Eagle could also be used for sex identification in Crested Goshawk. In this study, we have identified novel sex-specific DNA markers of Crested Serpent Eagle, Collared Scops Owl and Crested Goshawk. The use of these sex specific DNA markers would help in correctly identifying sexes before matching pair in a captive environment.