The Length Polymorphism of the 9th Intron in the Avian CHD1 Gene Allows Sex Determination in Some Species of Palaeognathae

Abstract

In palaeognathous birds, several PCR-based methods and a range of genes and unknown genomic regions have been studied for the determination of sex. Many of these methods have proven to be unreliable, complex, expensive, and time-consuming. Even the most widely used PCR markers for sex typing in birds, the selected introns of the highly conserved CHD1 gene (primers P2/P8, 1237L/1272H, and 2550F/2718R), have rarely been effective in palaeognathous birds. In this study we used eight species of Palaeognathae to test three PCR markers: CHD1i9 (CHD1 gene intron 9) and NIPBLi16 (NIPBL gene intron 16) that performed properly as Psittaciformes sex differentiation markers, but have not yet been tested in Palaeognathae, as well as the CHD1iA intron (CHD1 gene intron 16), which so far has not been used effectively to sex palaeognathous birds. The results of our research indicate that the CHD1i9 marker effectively differentiates sex in four of the eight species we studied. In Rhea americana, Eudromia elegans, and Tinamus solitarius, the electrophoretic patterns of the amplicons obtained clearly indicate the sex of tested individuals, whereas in Crypturellus tataupa, sexing is possible based on poorly visible female specific bands. Additionally, we present and discuss the results of our in silico investigation on the applicability of CHD1i9 to sex other Palaeognathae that were not tested in this study.

The development rule of feathers and application of hair root tissue in sex identification of Yangzhou geese

“…Specific primers can be designed for PCR amplification and the amplified products can be electrophoresed directly. The sex of the sample can be distinguished based on the number of bands generated; that is individuals with a single band are male ( ZZ ) and those with 2 bands are female ( ZW ) ( Kroczak et al, 2022 ). Here, this method was used to study sex identification in young geese.…”

Fast and Simple Molecular Test for Sex Determination of the Monomorphic Eudromia elegans Individuals

“…The vent sexing method is usually in concordance with the results of the molecular technique’s determination, except in two birds. Therefore, there is a small possible space for errors in such a method [ 2 , 15 ].…”

“…The development and optimization of molecular techniques for sex determination are crucial for both conservation and breeding programs. The method we employed builds upon the work of previous studies, enhancing the accuracy and reliability of sex determination in E. elegans [ 2 , 15 , 18 ]. Furthermore, the simplicity and cost-effectiveness of our PCR-based procedure can be beneficial for studies involving other avian species, especially those with limited genomic resources or in field settings.…”

“…The Paleognathaes form a group of less than 1% of all the extant avian species. The mostly terrestrial pheasant-like Tinamiformes species (Palaeognathae), including Eudromia elegans , inhabit the tropical New World biogeographic region that extends south, east, and west from the central plateau of Mexico [ 1 , 2 , 3 , 4 ].…”

Appropriate Primer Selection Improves Molecular Bird Sexing Accuracy

“…The PCR technique in birds sexing is based on the detection of the target gene chromodomain helicase DNA binding 1 (CHD1) on the Z and W chromosomes. It is known the sex chromosomes of male birds are composed of Z homogametes, whereas in female birds ZW heterogametes (Dobreva et al, 2021;England et al, 2021;Kroczak et al, 2022). Although the PCR method provides good results in determining the sex of birds, one of its successes is greatly influenced by the compatibility between the primers used and the DNA template (van der Velde et al, 2017).…”