Sexagem molecular em aves: abordagens genéticas e epigenéticas
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Data
2020-11-24
Autores
Jeronimo, Bruna Cristina [UNESP]
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Universidade Estadual Paulista (Unesp)
Resumo
Estima-se que aproximadamente 50% das espécies de aves não podem ter o sexo identificado por meio de características externas. A capacidade de identificar de maneira correta o sexo das aves é fundamental para o sucesso de estudos comportamentais, ecológicos, como também programas de reprodução em cativeiro com espécies ameaçadas de extinção. A identificação do gene CHD1 (Chromodomain Helicase DNA Binding Protein 1) possibilitou inúmeros avanços na sexagem molecular de aves. Este gene está presente nos cromossomos sexuais Z e W, as fêmeas possuem duas cópias diferentes, CHD1-Z e CHD1-W, enquanto os machos possuem duas cópias iguais do gene CHD1-Z. Recentemente, foi descrito um novo marcador associado a um loco que também apresenta homólogos nos cromossomos Z e W do gene Spindlin. Adicionalmente alguns autores também sugerem explorar diferenças epigenéticas entre machos e fêmeas com o intuito de identificar o sexo dos mesmos. No presente trabalho foram testados marcadores que exploram diferenças de tamanhos entre alelos dos genes CHD1 (P2/P8) e Spindlin (SPIN12/SPIN13), de genes específicos ao cromossomo W (ASW e XhoI) e também de um locus que apresenta diferenças de metilação de DNA entre machos e fêmeas de membros da superordem Galloanserae (MHM). Os marcadores relacionados ao cromossomo W não apresentaram resultados positivos, em contraposição os primers P2/P8 mostraram-se efetivos na sexagem molecular de indivíduos das ordens Piciformes e Psitaciformes, já amplicações com os marcadores SPIN12/SPIN13 apresentaram padrões sexo-específicos facilmente observáveis em géis de agarose em espécies das ordens Galliformes, Apodiformes e em exemplares de Corujinha-orelhuda. Nossos resultados demonstram que a escolha de um marcador molecular adequado para a identificação do sexo em aves está diretamente ligada a espécie em estudo e ao modo de visualização dos fragmentos gerados. Enquanto em algumas espécies os padrões sexo-especificos podem ser facilmente observáveis em agarose, outros dependem de métodos mais resolutivos, como poliacrilamida ou ainda, eletroforese capilar.
It is estimated that approximately 50% of bird species cannot have their sex identified by means of external characteristics. The ability to correctly identify the sex of birds is fundamental to the success of behavioral, ecological studies, as well as captive breeding programs with endangered species. The identification of the CHD1 gene (Chromodomain Helicase DNA Binding Protein 1) has enabled numerous advances in the molecular sexing of birds. This gene is located on the sex chromosomes Z and W, females have two different copies, CHD1-Z and CHD1-W, while males have two equal copies of the CHD1-Z gene. Recently, a new genetic marker associated with a locus that also has homologues copies on the Z and W chromosomes of the Spindlin gene has been described. Additionally, some authors also suggest exploring epigenetic differences between males and females in order to identify the sex. In the present work, molecular markers that explore size differences between alleles of the CHD1 (P2 / P8) and Spindlin (SPIN12 / SPIN13) genes, W-specific markers (ASW and XhoI) and also of a locus that presents epigenetic differences of DNA between males and females of members of the superorder Galloanserae (MHM) were tested. The molecular markers related to the W chromosome did not show positive results, in contrast, the P2 / P8 primers were effective in the molecular sexing of individuals of Piciform and Psitaciform orders, whereas amplifications with the SPIN12 / SPIN13 markers showed sex-specific patterns easily observed in agarose gels in species of the orders Galliformes, Apodiformes and stripd owl (Asio clamator). Our results demonstrate that the choice of a suitable molecular marker for the identification of sex in birds is directly linked to the species under study and to the method of visualization of the generated fragments. While in some species sex-specific patterns can be easily observed in agarose, others depend on more sensitive methods, such as polyacrylamide or capillary electrophoresis.
It is estimated that approximately 50% of bird species cannot have their sex identified by means of external characteristics. The ability to correctly identify the sex of birds is fundamental to the success of behavioral, ecological studies, as well as captive breeding programs with endangered species. The identification of the CHD1 gene (Chromodomain Helicase DNA Binding Protein 1) has enabled numerous advances in the molecular sexing of birds. This gene is located on the sex chromosomes Z and W, females have two different copies, CHD1-Z and CHD1-W, while males have two equal copies of the CHD1-Z gene. Recently, a new genetic marker associated with a locus that also has homologues copies on the Z and W chromosomes of the Spindlin gene has been described. Additionally, some authors also suggest exploring epigenetic differences between males and females in order to identify the sex. In the present work, molecular markers that explore size differences between alleles of the CHD1 (P2 / P8) and Spindlin (SPIN12 / SPIN13) genes, W-specific markers (ASW and XhoI) and also of a locus that presents epigenetic differences of DNA between males and females of members of the superorder Galloanserae (MHM) were tested. The molecular markers related to the W chromosome did not show positive results, in contrast, the P2 / P8 primers were effective in the molecular sexing of individuals of Piciform and Psitaciform orders, whereas amplifications with the SPIN12 / SPIN13 markers showed sex-specific patterns easily observed in agarose gels in species of the orders Galliformes, Apodiformes and stripd owl (Asio clamator). Our results demonstrate that the choice of a suitable molecular marker for the identification of sex in birds is directly linked to the species under study and to the method of visualization of the generated fragments. While in some species sex-specific patterns can be easily observed in agarose, others depend on more sensitive methods, such as polyacrylamide or capillary electrophoresis.
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Palavras-chave
Sexagem molecular, Aves, CHD, Spindlin, MHM, Análise para determinação do sexo, Epigenética, Sexo