Tree Genetics and Molecular Breeding 2024, Vol.14, No.4, 177-184 http://genbreedpublisher.com/index.php/tgmb 179 3 Advances in Genomic Tools for Dragon Fruit Research 3.1 Whole genome sequencing and annotation Whole genome sequencing has significantly advanced our understanding of dragon fruit genetics, particularly with the sequencing of Hylocereus undatus, the white-fleshed dragon fruit. This chromosomal-level genome assembly revealed approximately 29 000 protein-coding genes, similar to other members of the Cactaceae family, such as Carnegiea gigantea. The sequencing efforts have uncovered a whole-genome duplication event in the last common ancestor of Cactaceae, followed by extensive genome rearrangements (Salgotra and Stewart, 2020). This genomic information is crucial for identifying genes associated with important traits such as drought resistance and fruit flavor, which are overrepresented in the orthologous gene clusters of H. undatus (Zheng et al., 2021; Xu and Wang, 2024). 3.2 Transcriptomics and differential gene expression studies Transcriptomics has emerged as a powerful tool in dragon fruit research, enabling the study of gene expression patterns under various conditions. The availability of extensive OMICS datasets, including transcriptomics, has facilitated the identification of gene-to-trait associations, which are essential for understanding the genetic basis of performance traits in fruit trees. These studies help in pinpointing genes that are differentially expressed in response to environmental stresses or during different developmental stages, providing insights into the molecular mechanisms underlying these processes (Gouthu et al., 2022; Khokhar et al., 2023). 3.3 CRISPR-Cas9 applications in dragon fruit breeding The application of CRISPR-Cas9 technology in dragon fruit breeding holds great promise for precision breeding. This genome-editing tool allows for targeted modifications in the dragon fruit genome, enabling the development of new cultivars with desirable traits such as improved stress resistance and enhanced nutritional content (Rai and Shekhawat, 2015). Although the fruit tree research community has been slow to fully embrace these technologies compared to staple crops, the potential for CRISPR-Cas9 to revolutionize dragon fruit breeding is significant. The technology's ability to create transgene-free edits through Ribonucleoprotein (RNP) delivery is particularly appealing, as it aligns with public preferences for sustainable and eco-friendly agricultural practices (Iwata et al., 2016; Minamikawa et al., 2017). 4 Case Study 4.1 Floral development and reproductive challenges inH. monacanthus lines Since 2009, seedlings from five Hylocereus monacanthus androgenic lines have been cultivated under greenhouse conditions (Garcia et al., 2009a; Garcia et al., 2009b). These lines displayed suboptimal growth characteristics, including extremely slow vegetative development and an extended seedling stage lasting several years. Morphologically, the stems and spines of these plants resembled the donor species, but notable differences were observed in floral development. Flower buds in these androgenic lines showed early abscission prior to blooming. Among the lines, two (007 and 011) initiated flower development in October 2013, with the remaining three lines following suit a year later (Figure 1A; Figure 1B). During the early stages of floral development, all five lines exhibited flower bud morphology similar to the donor species, characterized by green sepals with a red-purple margin (Figure 1A). However, aberrations became evident in the upper floral structures, including flattened and folded apical ends with inward-folded sepals exposing stigmas. Both pistils and anthers exhibited atypical deformities (Figure 1B; Figure 1C). Most flower buds abscised before full bloom, preventing the assessment of pollen and ovule viability. Histological analysis of a 6 cm flower bud from line 007 revealed that the ovary contained significantly smaller, deformed, and shriveled ovules compared to the donor plant, which exhibited larger, typical plump ovules of spherical shape (Figure 1D; Figure1 E). Observations of anthers from line 007 further indicated marked degeneration, with the majority of pollen grains being sterile and exhibiting an abnormal triangular shape or wrinkled tapetal layers (Figure 1F). In contrast, the donor plant showed normal tetrads with a typical near-spherical shape (Figure 1G). Interestingly, line 011 successfully developed a single flower to full bloom in October 2016 (Figure 1H) and
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