Tree Genetics and Molecular Breeding 2024, Vol.14, No.4, 194-205 http://genbreedpublisher.com/index.php/tgmb 200 RNA sequence to introduce double-strand breaks at target sites, which are then repaired by cellular mechanisms, leading to desired genetic modifications (Sander and Joung 2014; Zhang et al., 2014). In the context of sex determination studies, CRISPR/Cas9 can be utilized to knock out or modify genes implicated in sex differentiation pathways. For instance, in Eucommia ulmoides, key genes such as EuAP3 and EuAG have been identified as regulators of sex differentiation, and CRISPR/Cas9 could be employed to elucidate their specific roles by creating targeted mutations. Despite its potential, the application of CRISPR/Cas9 in sex determination studies must address challenges such as off-target effects and mosaicism, which can complicate the interpretation of results (Liang et al., 2016). Figure 2 Y-chromosomal fitness effects alter the scope for SD transitions (Adopted from Schenkel et al., 2022) Image caption: Shown are the predicted equilibrium frequencies of FI in females (maternally inherited alleles), MY and MA inmales (paternally inherited alleles); we restrict our analysis to the maternal (FI) c.q. paternal (MY, MA) alleles to account for their (potential) sex-specific transmission. Horizontal labels indicate locus and temperature, vertical labels the MA activation rate μD. Further parameter values used in simulations: θF=1.2; θM=0.3; β=0.5. Predicted allele frequencies were smoothed with binomial generalized additive models (Adopted from Schenkel et al., 2022) 6.2 Development of sex-specific markers and their application in breeding The development of sex-specific markers is crucial for the efficient breeding of dioecious plants like Eucommia ulmoides. These markers can be used to identify the sex of plants at an early stage, thereby facilitating selective breeding and cultivation practices. Advances in genomic technologies, such as high-throughput sequencing and bioinformatics, have enabled the identification of sex-linked markers. For example, the chromosome-level genome assembly of E. ulmoides has provided valuable insights into the genetic basis of sex differentiation, allowing researchers to pinpoint specific markers associated with male and female traits (Belhaj et al., 2015). These markers can be applied in breeding programs to ensure the propagation of desired sex types, thereby optimizing yield and resource allocation.
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