Medicinal Plant Research 2024, Vol.14, No.2, 71-84 http://hortherbpublisher.com/index.php/mpr 74 “QX-145” (mother) and “Nannong Yinshan” (father), containing 109 and 125 marker loci, respectively, and comprising 25 and 21 linkage groups. The cumulative map lengths were 1,465.6 cM and 1,972.7 cM. These early works laid a theoretical foundation for deciphering the genetic structure of Chrysanthemum morifolium and accelerated the development of marker-assisted breeding programs (Zhang et al., 2010). The advent of high-throughput sequencing technology has taken Chrysanthemum genomics research to a new level, providing crucial references for understanding the phenotypic characteristics and the mechanisms of medicinal compound synthesis in Chrysanthemum morifolium(Liu et al., 2015; Sasaki et al., 2017). In 2017, the Chrysanthemum Whole Genome Project, jointly initiated by Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences and the Amway Plant Research and Development Center, made significant progress. The joint team utilized nanopore sequencing technology to overcome the challenges of sequencing complex higher plant genomes, completing the whole genome sequencing of the medicinal Chrysanthemum species for the first time. They also accomplished the full-length transcriptome genetic information extraction of Hangbaiju (Chrysanthemum morifolium 'Hangbaiju'). Subsequently, the genomes of Chrysanthemum seticuspe and Chrysanthemum nankingense were also assembled (Song et al., 2018; Hirakawa et al., 2019). In 2023, the team led by Chen Fadi from Nanjing Agricultural University decoded the genome of the hexaploid cultivated Chrysanthemum morifolium, reporting the world's first segmental allopolyploid genome. This research delved into the origin and breeding history of cultivated Chrysanthemum morifolium, laying a foundation for the next steps in deciphering the molecular mechanisms behind important horticultural traits (such as flower shape, flower color, plant form, and stress resistance) and targeted breeding. The release of the Chrysanthemum genome facilitates the in-depth study of high-density genetic map construction and advances the research in the biology and molecular breeding of flowering plants (Hirakawa et al., 2019). 3.2 Methods and technologies used in genome sequencing The methodologies employed in the genomic studies of Chrysanthemum morifolium have evolved with technological advancements. Early studies utilized molecular markers such as RAPD, ISSR, and AFLP to construct genetic linkage maps (Zhang et al., 2010), but these maps had relatively large average genetic distances between markers and contained fewer markers. The rapid development of high-throughput sequencing technologies, including whole genome sequencing, reduced representation genome sequencing, and RNA-Seq, has provided more cost-effective and efficient means for molecular marker development. First-generation sequencing technologies primarily included the Sanger method of chain-termination sequencing and the chemical degradation method (Sanger et al., 1977). The main advantages of first-generation sequencing were long read lengths and high base reading accuracy, but these methods also had significant drawbacks of low throughput and high cost. Second-generation sequencing technologies, also known as next-generation sequencing (NGS) or deep sequencing (Sultan et al., 2008), include four main sequencing technologies developed over time: pyrosequencing by Roche 454, Solexa sequencing by Illumina, Supported Oligo Ligation Detection (SOLiD) sequencing by ABI, and DNBSEQ sequencing by BGI. Among these, Roche 454's pyrosequencing and ABI's SOLiD have been phased out. Second-generation sequencing technologies have facilitated the acquisition of large-scale transcriptome data and EST markers, promoting the identification of genes related to various traits (Sasaki et al., 2017; Yue et al., 2018). Unlike the SMRT sequencing technology by PacBio, which converts biochemical signals into fluorescence signals for sequencing, Oxford Nanopore Technology's nanopore sequencing technology uses electrical signals for sequencing. Some classify this new nanopore sequencing technology as fourth-generation sequencing (Feng et al., 2015), but due to its long read length characteristic, it is considered third-generation sequencing in this context. Compared to second-generation sequencing, third-generation single-molecule sequencing technology offers significant advantages such as long read lengths, fast sequencing speed, and the ability to sequence complex genomic regions. This makes it indispensable for assembling complex genomes, detecting large structural
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