Triticeae Genomics and Genetics, 2024, Vol.15, No.2, 66-76 http://cropscipublisher.com/index.php/tgg 67 2 Molecular Tools inTriticeae 2.1 Genomic sequencing technologies Recent advancements in next-generation sequencing (NGS) technologies have significantly reduced the cost of DNA sequencing, enabling the sequencing of species with large and complex genomes, such as bread wheat (Triticum aestivum L.) (Hussain et al., 2022). The international effort to sequence the wheat genome has culminated in the release of a fully annotated reference wheat-genome assembly, which has paved the way for the pan-genomic era in wheat research. This has facilitated the use of genotyping arrays capable of characterizing hundreds of wheat lines with thousands of markers, providing fast, relatively inexpensive, and reliable data for wheat breeding (Hussain et al., 2022). Additionally, open-source computational workflows like TRITEX have been developed to construct chromosome-scale sequence assemblies, further enhancing genomic studies in Triticeae crops (Monat et al., 2019). 2.2 Marker-assisted selection (MAS) Marker-assisted selection (MAS) has revolutionized plant breeding by improving the productivity and accuracy of classical breeding methods. DNA markers have been extensively used to map quantitative trait loci (QTLs) and associate them with specific genes, thereby accelerating the development of new crop varieties (Hasan et al., 2021). Functional markers (FMs) closely associated with phenotypic traits have been identified using various genomics approaches, facilitating the direct selection of genes associated with desirable traits (Salgotra and Stewart, 2020). MAS has been successfully applied in breeding programs to improve traits such as drought resilience in alfalfa, demonstrating its potential to enhance crop productivity under stress conditions (Singh et al., 2022). 2.3 CRISPR-Cas9 and genome editing CRISPR-Cas9 has emerged as a powerful, cost-effective, and versatile tool for precise and efficient genome editing in plants. Unlike earlier genome editing tools like zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs), CRISPR-Cas9 involves simpler designing and cloning methods, making it more accessible for targeted mutagenesis (Jaganathan et al., 2018; Razzaq et al., 2019). This technology has been used for single base substitution, multiplex gene editing, gene knockouts, and regulation of gene transcription, significantly contributing to crop improvement (Razzaq et al., 2019). The development of CRISPR-Cas12a and other advanced CRISPR systems has further enhanced the efficiency and specificity of genome editing, enabling the engineering of multiple genes simultaneously (Liu et al., 2019). These advancements promise to bridge the gap between forward and reverse genetics, providing rapid methods to validate genes and alleles for crop improvement (Thomson et al., 2022). 2.4 Transcriptomics and gene expression profiling Functional genomics approaches, including transcriptomics, have been instrumental in identifying genes and functional markers associated with plant phenotypic variation. Transcriptomics involves the comprehensive analysis of gene expression profiles, which helps in understanding the molecular mechanisms underlying various traits (Salgotra and Stewart, 2020). This approach, combined with other functional genomics strategies like TILLING and association mapping, has been used to identify candidate genes linked to key traits such as grain yield, end-use quality, and resistance to biotic and abiotic stresses in wheat (Hussain et al., 2022). The integration of transcriptomics with other molecular tools provides a holistic view of gene function and regulation, facilitating the development of improved crop varieties. In summary, the integration of genomic sequencing technologies, marker-assisted selection, CRISPR-Cas9 genome editing, and transcriptomics has significantly advanced the molecular breeding of Triticeae crops. These molecular tools offer new opportunities to enhance crop productivity and resilience, addressing the challenges of global food security.
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