Bioscience Methods 2024, Vol.15, No.6, 289-301 http://bioscipublisher.com/index.php/bm 298 9 Challenges and Future Directions in Rubber Research 9.1 Technical challenges in biosynthesis pathway modification Modifying the biosynthesis pathways of rubber in both Eucommia ulmoides and Hevea brasiliensis presents several technical challenges. One significant challenge is the complexity of the rubber biosynthesis pathways themselves. In E. ulmoides, the methylerythritol-phosphate (MEP) pathway is predominantly used for isoprenyl diphosphate synthesis, whereas H. brasiliensis primarily utilizes the mevalonate (MVA) pathway (Chow et al., 2011; Li et al., 2020). This difference necessitates distinct approaches for pathway modification in each species. Additionally, the regulation of these pathways involves numerous genes and enzymes, making targeted modifications difficult. For instance, the role of long non-coding RNAs (lncRNAs) in regulating rubber biosynthesis in E. ulmoides adds another layer of complexity (Liu et al., 2018). Furthermore, the presence of transposable elements (TEs) and their derived small interfering RNAs (siRNAs) in H. brasiliensis can interfere with gene expression, complicating genetic modifications (Wu et al., 2020). 9.2 Opportunities for genetic engineering Despite these challenges, there are significant opportunities for genetic engineering to enhance rubber production. Advances in genome sequencing and assembly have provided high-quality genomic resources for both E. ulmoides and H. brasiliensis, facilitating the identification of key genes involved in rubber biosynthesis (Lau et al., 2016; Wuyun et al., 2017; Li et al., 2020). Genetic engineering can target these genes to improve rubber yield and quality. For example, the expansion of rubber biosynthesis-related gene families in H. brasiliensis suggests potential targets for enhancing latex production (Lau et al., 2016). Additionally, the identification of differentially expressed lncRNAs and microRNAs in H. brasiliensis offers new avenues for manipulating gene expression to increase rubber yield (Li et al., 2022). The development of transgenic rubber trees with enhanced biosynthetic pathways could significantly boost rubber production (Chow et al., 2011). 9.3 Future research directions in rubber biosynthesis Future research in rubber biosynthesis should focus on several key areas. First, a deeper understanding of the regulatory networks controlling rubber biosynthesis is essential. This includes studying the roles of lncRNAs, microRNAs, and TEs in gene regulation (Liu et al., 2018; Wu et al., 2020; Li et al., 2022). Second, research should aim to elucidate the final stages of rubber elongation, which remain poorly understood (Wu et al., 2020). Third, comparative studies between E. ulmoides and H. brasiliensis can provide insights into the evolution of rubber biosynthesis pathways and identify common regulatory mechanisms (Wuyun et al., 2017). Finally, integrating genomic, transcriptomic, and epigenetic data will be crucial for developing effective genetic engineering strategies to enhance rubber production (Chow et al., 2007; Lau et al., 2016; Li et al., 2020). By addressing these research directions, we can improve our understanding of rubber biosynthesis and develop innovative approaches to meet the growing demand for natural rubber. 10 Concluding Remarks This comparative study of rubber biosynthesis pathways in Eucommia ulmoides and Hevea brasiliensis has revealed significant differences and similarities in their genetic and biochemical mechanisms. The high-quality genome assembly of E. ulmoides has provided new insights into its evolution and rubber biosynthesis, highlighting the reliance on the methylerythritol-phosphate (MEP) pathway for isoprenyl diphosphate synthesis, which is predominantly active in trans-polyisoprene-containing leaves and central peels. In contrast, H. brasiliensis primarily utilizes the mevalonate (MVA) pathway for cis-polyisoprene biosynthesis, with evidence suggesting a potential role for the MEP pathway as well. Additionally, the expansion of gene families related to rubber biosynthesis in H. brasiliensis has been identified as a key factor contributing to its high latex yield. Differential expression of long noncoding RNAs (lncRNAs) and microRNAs between self-rooting juvenile clones and donor clones of H. brasiliensis further unveils the molecular regulation underlying increased rubber yield. The insights gained from this study have several potential applications. The high-quality genome assembly of E. ulmoides can facilitate genetic engineering efforts to enhance its industrial and medicinal uses. Understanding the distinct biosynthesis pathways in E. ulmoides and H. brasiliensis can lead to the development of transgenic rubber
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