Oil palm (Elaeis guineensis Jacq.) is one of the most important tropical oil crops in the world. With the growing global population, ensuring the sustainable development of the palm oil industry is crucial to meet the increasing demand for palm oil consumption (Wang et al. 2019). However, the sustainability and productivity of oil palm plantations, crucial for the global vegetable oil industry, face significant challenges. The growth in yield is hampered by shrinking planted areas and reduced arable land. Several factors, such as crop loss to pests and diseases, impact productivity. Unpredictable weather patterns due to climate change affect fruiting cycles and overall productivity. Maintaining soil fertility and moisture content, especially in peatland areas, remains a persistent challenge, resulting in substantial yield losses (Parveez et al. 2023). The use of tissue culture to produce oil palm clones enables rapid multiplication of genetically superior palms while maintaining their desirable traits. Tissue culture accelerates propagation, preserves genetic purity, and ensures uniform and predictable plantations. Furthermore, it integrates genetic engineering and molecular breeding, enhancing the overall genetic quality of oil palms. The challenges faced by oil palm plantations demand a comprehensive approach that combines traditional breeding with modern biotechnological advancements, essential for developing varieties with desired traits and ensuring the industry's long-term sustainability (Low et al. 2008; Sahara et al. 2023).
Somatic embryogenesis (SE) is an effective technique for the mass plant production, particularly in perennial crops like oil palm (Zhang et al. 2021). Despite its importance, a notable obstacle in oil palm tissue culture lies in the restricted rate of somatic embryogenesis (Ooi et al. 2021; Sahara et al. 2023). The successful development of somatic embryos relies on efficient embryogenic callus induction, making it a critical step for subsequent somatic embryo development. In the aspect of improving phenotypic traits through genetic engineering, it is important for the callus derived from recalcitrant plant species to be generable to intact plant (Tuskan et al. 2018).
Recent research conducted by (Ong-Abdullah and Siew Eng 2007) has identified key genes, such as EgPER1, EgHOX1, and EgPK1, that distinguish between embryogenic and non-embryogenic calli in oil palm tissue culture. In a study by (Low et al. 2008), candidate genes were pinpointed as being differentially expressed during callogenesis and embryogenesis in oil palm. These genes include lipid transfer proteins, metallothionein-like proteins, glutathione S-transferase, and dehydrin-like protein. Ooi et al. (2021) proposed in their article that genes linked to flowering time, light response, photosynthesis, and stress response may be correlated with somatic embryogenesis potential in oil palm. Sahara et al. (2023) reported the transcriptome profiling of high- and low-embryogenic ortets of oil palm at the callus and somatic embryoid stages using RNA-seq. This article contributes potential differentially expressed genes (DEGs) for high-embryogenic ortets, thereby enhancing the efficiency of oil palm micropropagation. Recent research on maize has highlighted the genotype-dependent nature of embryogenic callus induction (Liang et al. 2023). Several genes responsible for callus induction have been identified in numerous plant species. For instance, the WAK gene family in Chinese cabbage has shown potential involvement in callus cell growth and reproduction (Zhang et al. 2020a). In Arabidopsis, LATERAL ORGAN BOUNDARIES DOMAIN (LBD) genes play a key role in callus induction (Fan et al. 2012a). The AtbZIP59–LBD complex is crucial in regulating auxin-induced callus formation (Xu et al. 2018), while CYCLIN D3 (CYCD3) is involved in wounding-induced callus formation (Ikeuchi et al. 2017). Moreover, NAC DOMAIN CONTAINING PROTEIN71 (ANAC071) and AP2/ERF transcription factor RAP2.6L have been identified as essential regulators during wound-induced callus formation process in Arabidopsis (Ikeuchi et al. 2013). Studies in cotton have indicated that the WOX genes play a pivotal role in callus induction (Muhammad Tajo et al. 2022). Overexpression of ZmBBM2 promotes callus formation (Du et al. 2019), and ZmMYB138 transcription factor being another promoter of callus formation via GA signal transduction in maize (Ge et al. 2016). Additionally, ZmARF23 mediates callus induction by binding to the ZmSAUR15 promoter and enhancing its transcriptional expression (Liang et al. 2023). In Panax ginseng, silencing the PgWRKY6 gene reduced the rate of embryogenic callus induction (Yang et al. 2020). As genetic factors play a crucial role in callus induction and proliferation, identifying genes and regulatory elements responsible for controlling callus formation can provide valuable insights into developing in vitro systems for recalcitrant plant species especially in oil palm (Tuskan et al. 2018; Weckx et al. 2019).
Genome-wide association study (GWAS) has emerged as an effective tool to investigate the associations between genotypes and phenotypes, facilitating the identification of single nucleotide polymorphisms (SNPs) linked to phenotypic variation within a population (Alqudah et al. 2020). GWAS have been conducted in oil palm to identify markers and candidate genes associated with various traits. These studies have used genotyping-by-sequencing (GBS) and SNP analysis to identify SNPs that are significantly associated with traits such as height (Babu et al. 2019b; Somyong et al. 2022), morphological and yield-related traits (Osorio-Guarín et al. 2019), fatty acid composition (Xia et al. 2019), and leaf area, rachis length and total dry weight (Babu et al. 2019a). These findings have the potential to improve the efficiency and effectiveness of oil palm breeding programs through marker-assisted selection (MAS) and targeted functional analyses. The method's adaptability across different plant species, including oil palm (Babu et al. 2019b, a; Xia et al. 2019; Osorio-Guarín et al. 2019; Somyong et al. 2022), rice (Yano et al. 2016; Zhang et al. 2019b), rapeseed (Qu et al. 2017), rose (Nguyen et al. 2020), Populus (Tuskan et al. 2018; Zhang et al. 2020b), maize (Guo et al. 2020; Shikha et al. 2021), sesame (Cui et al. 2021), Gossypium arboretum (Zhao et al. 2021; Hu et al. 2022), Senegalese sorghum (Ahn et al. 2023), and wheat (Ma et al. 2023; Gudi et al. 2023), demonstrates its versatility and broad applicability in crop improvement and breeding programs.
A GWAS identifies clusters of linked SNPs associated with the target trait, and the development of molecular markers linked to a trait of interest facilitates MAS during the early stages of plant development (Rahman et al. 2007; Uffelmann et al. 2021). Extensive studies have successfully detected SNP markers for selecting desired traits (Hayashi et al. 2004; Sattarzadeh et al. 2006; Gaudet et al. 2007; Kim et al. 2022), supporting the effectiveness of MAS. However, genomic loci and effective markers related to callus induction in oil palm remains unclear and related studies is required.
In this study, callus induction rates at 1-, 2-, and 3-months after inoculation (C1, C2 and C3) were observed in immature male inflorescences of 198 oil palm accessions for phenotyping, along with genotyping by resequencing. GWAS was then conducted accordingly to identify SNP loci linked to callus induction rates. Subsequently, a specialized SNP marker, designed to distinguish alleles with a high potential for callus induction, was developed to facilitate the process of ortet palm selection at early stages. This strategic marker not only expedites the identification of promising ortets but also addresses challenges related to the efficient and accurate selection of ortet palm. The issues encountered by the selection of ortet palms, such as variability in callus induction potential and genotypic complexities, are mitigated by the precision and discriminatory power of the developed SNP marker. The utilization of this SNP marker in the early stages allows for rapid and targeted identification of ortets with high potential of callus induction, significantly enhancing the efficiency of oil palm breeding programs.