Development of Y‐chromosome‐Specific SCAR Markers Conserved in Taurine,Zebu and Bubaline Cattle |
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| Authors: | BCA Alves VFM Hossepian de Lima CA Moreira‐Filho |
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| Institution: | 1. Biotechnology Research Center, University of S?o Paulo;2. Present address: Albert Einstein Research and Education Institute, Av. Albert Einstein, 627,CEP: 05651‐901, S?o Paulo, SP, Brazil;3. Department of Animal Reproduction, School of Veterinary and Agronomical Sciences, UNESP, Jaboticabal;4. Department of Pediatrics, University of S?o Paulo School of Medicine, S?o Paulo, SP, Brazil |
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| Abstract: | Sex pre‐selection of bovine offsprings has commercial relevance for cattle breeders and several methods have been used for embryo sex determination. Polymerase chain reaction (PCR) has proven to be a reliable procedure for accomplishing embryo sexing. To date, most of the PCR‐specific primers are derived from the few single‐copy Y‐chromosome‐specific gene sequences already identified in bovines. Their detection demands higher amounts of embryonic genomic material or a nested amplification reaction. In order to circumvent this, limitation we searched for new male‐specific sequences potentially useful in embryo sexing using random amplified polymorphic DNA (RAPD) analysis. Random amplified polymorphic DNA (RAPD) assay reproducibility problems can be overcome by its conversion into Sequence Characterized Amplified Region (SCAR) markers. In this work, we describe the identification of two bovine male‐specific markers (OPC16323 and OPF101168) by means of RAPD. These markers were successfully converted into SCARs (OPC16726 and OPF10984) using two pairs of specific primers.Furthermore, inverse PCR (iPCR) methodology was successfully applied to elongate OPC16323 marker in 159% (from 323 to 837 bp). Both markers are shown to be highly conserved (similarity ≥95%) among bovine zebu and taurine cattle; OPC16323 is also highly similar to a bubaline Y‐chromosome‐specific sequence. The primers derived from the two Y‐chromosome‐specific conserved sequences described in this article showed 100% accuracy when used for identifying male and female bovine genomic DNA, thereby proving their potential usefulness for bovine embryo sexing. |
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