Applications of Plant Transformation

Introduction

Plant transformation technology involves the introduction of foreign genes into the plant genome, enabling targeted modification of plant genetic traits, thereby promoting agricultural modernization and biotechnology development. This technology not only provides new tools for crop improvement but also demonstrates broad potential for applications in areas such as nutritional enhancement, stress tolerance enhancement, and pharmaceutical compound production. With the advancement of gene-editing technologies such as CRISPR-Cas9, plant transformation has evolved from traditional random integration to precise regulation, becoming a key tool for addressing global food security and environmental challenges.

Overview of Plant Transformation Technology

Plant transformation technology is a core means of genetic improvement by introducing exogenous genes into the plant genome. Various methods exist, primarily including the following:

Agrobacterium-mediated Transformation

Agrobacterium tumefaciens mediates the stable integration of exogenous genes via the T-DNA region of the Ti plasmid. By modifying the T-DNA of the Ti plasmid, the target gene can be integrated into the plant genome, allowing plants to be regenerated through tissue culture. Agrobacterium-mediated transformation is simple and efficient, and has been widely used in dicotyledons (such as tobacco and tomato). However, monocotyledons (such as corn and wheat) lack actively dividing cells, resulting in lower transformation efficiency. Therefore, optimization of the explant type (such as immature embryos) and Agrobacterium strain (such as highly virulent strains) is necessary to increase transformation success rates.

Gene Gun Technique

Gene gun technique uses high-pressure gas to accelerate metal microparticles (such as gold powder) coated with the target gene through plant cell membranes, achieving gene delivery. This method is applicable to a wide range of plants, but suffers from low transformation efficiency, random integration of the exogenous DNA, and poor genetic stability.

Direct DNA Transfer Techniques

• Electroporation: This technique uses high-voltage electric pulses to create pores in the cell membrane, facilitating DNA entry into cells. Electroporation is applicable to both monocotyledons and dicotyledons, but requires optimization of electric field parameters and DNA concentration to improve transformation efficiency.
• Liposome-mediated method: This technique uses liposomes to encapsulate DNA, then transfers the gene through cell membrane fusion. Liposome-mediated technology is commonly used for somatic cell transformation.
• CRISPR/Cas system: This gene-editing technology allows for precise knockout or insertion of specific genes, but CRISPR/Cas system has not yet been widely adopted in commercial crops.

Comparison of Stable and Transient Transformation

• Stable transformation refers to the integration of a foreign gene into the plant genome and its stable inheritance. Stable transformation requires regeneration of intact plants through tissue culture to ensure stable inheritance and expression of the foreign gene. It is suitable for long-term trait improvement (such as insect resistance and stress tolerance). For example, the introduction of the EPSPS gene into soybeans through Agrobacterium-mediated transformation confers herbicide resistance.
• Transient transformation refers to the transient expression of the foreign gene in the cell nucleus. Transient transformation directly introduces single-stranded DNA into the cell nucleus without integrating it into the genome. This method is suitable for short-term functional studies (such as promoter activity analysis). Transient transformation allows for rapid screening of candidate genes, but it does not produce stably inherited transgenic plants.

Key Influencing Factors

• Transformation efficiency: This is influenced by the state of the recipient material, the Agrobacterium strain, and treatment conditions. For example, Arabidopsis thaliana achieved the higher transformation efficiency when treated with floral dip treatment during the young flowering stage, while biolistic methods require optimization of DNA concentration and targeting to improve efficiency.
• Genotype dependence: Different plants exhibit significant differences in their response to transformation methods. Dicots are often treated with Agrobacterium, while monocots rely on biolistic methods or electroporation.
• Selection strategies: Antibiotic resistance selection (e.g., kanamycin, hygromycin) or reporter gene labeling (e.g., GUS, LUC) for transformed cells are commonly used.

Genetic Improvement through Plant Transformation

Plant transformation technology imparts traits such as stress tolerance, improved nutrition, and herbicide resistance to crops by stably integrating foreign genes into the plant genome. Specific application areas include:

Biotic Stress Resistance

• Insect resistance: The toxin protein encoded by the Bt gene (from Bacillus thuringiensis) damages the insect gut epithelial cells, conferring resistance to pests such as cotton bollworm and corn borer. For example, Bt cotton was successfully commercialized through Agrobacterium-mediated transformation and has become a major insect-resistant crop worldwide.
• Viral resistance: Virus unpacking can be blocked by using coat protein genes, or viral replication can be inhibited through antisense RNA and ribozyme-mediated strategies. For example, virus-resistant tomatoes achieve resistance by expressing viral coat protein genes.
• Disease resistance: Resistance to specific pathogens can be conferred by using R genes (such as Rps1k). For example, transgenic eggplants express R genes to resist bacterial wilt.
• Synergistic multi-stress resistance: By simultaneously introducing multiple resistance genes (e.g., Bt + R genes), a composite resistance system is constructed to enhance crop defense against multiple pathogens.

Abiotic Stress Tolerance

• Drought resistance: Overexpression of transcription factors (e.g., DREB, HSP) or osmoprotectants (e.g., proline). For example, transgenic rice (IR64-DREB) maintains photosynthetic efficiency under drought conditions.
• Salt tolerance: Overexpression of ion transporters (e.g., NHX1) regulates intracellular ion balance. For example, transgenic rapeseed improves salt tolerance through expression of the NHX1 gene.
• Cold tolerance: Introducing antifreeze protein genes (e.g., antifreeze protein, AFP) enhances cold stress tolerance. For example, transgenic wheat prolongs cold survival through expression of AFP genes.

Fig. 1. Major abiotic stresses and biotechnological tools to help plants to resist these stress conditions. (Imran, et al. 2021)

Nutrition And Metabolic Engineering

• Biofortification: Increasing vitamin A content through multi-gene integration. For example, Golden Rice enriches its endosperm with β-carotene by overexpressing β-carotene synthase genes (e.g., LCYB and LCYE).
• Oil and fat modification: Modifying oil and fat composition by manipulating the fatty acid biosynthesis pathway (e.g., FAD3 and FAD2 genes). For example, transgenic soybeans overexpress the Δ6-FAD gene to increase oleic acid content.
• Iron enhancement: Increasing iron bioavailability through the expression of ferritin or metallothionein genes. For example, transgenic rice expresses ferritin genes to enhance iron absorption.

Herbicide Resistance

• Commonly used genes: EPSPS (5-enolpyruvic acid manganate-3-phosphate synthase) and BAR (bialaphos resistance gene) confer resistance to glyphosate and phosphinothricin. For example, transgenic soybeans overexpress the EPSPS gene to tolerate high doses of glyphosate.
• Management impacts: Herbicide-tolerant crops can reduce farmers' reliance on herbicides and reduce environmental pollution. For example, herbicide-tolerant maize (MON810) increases crop yield by reducing weed competition.

Crop Improvement and Trait Customization

Yield Improvement

Through genetic engineering, scientists can precisely manipulate a plant's flowering time, plant architecture, and sink-source relationships, thereby optimizing light energy utilization efficiency and photosynthate distribution. For example, introducing genes that control plant architecture (such as dwarfing genes) can reduce ineffective tillering and increase yield per unit area. Furthermore, the introduction of growth regulators (such as gibberellins or cytokinins) can further promote robust plant growth. In practical applications, insect-resistant corn (such as Bt corn) significantly reduces pest damage by expressing Bacillus thuringiensis toxin genes, indirectly increasing yield. Gene editing technologies (such as CRISPR-Cas9) can also target yield-related quantitative trait loci (QTLs), directly introducing high-yield alleles and accelerating breeding efforts.

Quality Trait Improvement

• Flavor and appearance: By introducing genes related to anthocyanin metabolism (such as the MYB transcription factor), crops with novel flower colors (such as pink tomatoes) can be bred. In addition, overexpressing specific genes (such as the β-carotene synthase gene) can enhance the nutritional value of crops (e.g., the vitamin A content in Golden Rice).
• Stress tolerance: Gene editing technology can be used to selectively knock out or overexpress genes related to fruit ripening (such as the ACO and PG genes), delaying ripening and extending the shelf life of tomatoes. Similarly, introducing drought-resistant genes (such as FvC5SD) can enhance drought tolerance in crops.
• Seedless fruit transformation and disease resistance: Seedless fruit has been achieved in tomatoes (e.g., by suppressing genes related to seed development), while disease resistance has been improved by introducing antiviral genes (such as viral capsid protein genes) or insect-resistant genes (such as the Bt gene).

Post-Harvest Improvement

• Storability: Gene editing (e.g., knocking out α-mannosidase and β-hexosidase) can significantly extend the shelf life of tomatoes and peppers. Furthermore, genetic modification can reduce antinutrients (such as oxalic acid) in crops, improving food safety.
• Cold chain durability: Regulating freeze-responsive genes (such as antifreeze protein genes) can enhance crop stability in low-temperature environments.
• Disease resistance: Transgenic potatoes expressing the Amaranthus hypochondriacus seed protein AmA1 exhibit enhanced disease resistance.

Hybrid Seed Production and Male Sterility

• Cytoplasmic male sterility (CMS): Introducing CMS genes into crops through transgenic technology enables efficient hybrid seed production and reduces self-interference.
• Barnase/barstar system: This system, which expresses the barnase enzyme to inactivate pollen viability while simultaneously protecting the female parent from sterility through the barstar inhibitor, is widely used in rice and corn hybrids.
• Controversy and challenges: Although terminator genes (such as patatin) can induce seed sterility, their commercial application is limited by ecological risks and ethical concerns.

Modern Transgenic Applications in Precision Agriculture

CRISPR-enabled Trait Engineering

Gene-editing technologies such as CRISPR-Cas9 enable targeted improvement of crop traits by precisely targeting genomic loci. For example, CRISPR-Cas9 has been successfully used to improve disease resistance in rice (e.g., knocking out genes associated with rice blast) and enhance drought resistance in wheat. Compared to traditional transgenics, gene-editing technologies do not require the introduction of foreign DNA, reducing regulatory risks. Furthermore, multi-gene editing (e.g., simultaneous editing of multiple resistance genes) allows for the rapid development of multi-resistance varieties.

Synthetic Biology and Metabolic Pathway Design

Synthetic biology approaches can be used to engineer crop secondary metabolic pathways to produce high-value-added compounds. For example, gene editing has been used to overexpress the β-carotene synthesis gene in rice, resulting in the development of vitamin A-rich golden rice. Furthermore, the design of novel biosynthetic pathways (e.g., engineering plants to produce antibiotics or industrial enzymes) can provide sustainable raw materials for the pharmaceutical and industrial sectors.

Fig. 2. CRISPR-mediated plant synthetic biology in which plant cell behavior is altered to enhance plant growth and product generation. (Gao, C. 2021)

Non-GM Genome Editing Methods

• RNP complex delivery: Transient expression of the Cas9 protein and sgRNA enables gene editing without DNA integration, avoiding the linkage drag issues of traditional transgenics.
• Site-specific recombination (SSR): Homologous recombination techniques can efficiently repair targeted sites, generating crops without transgenic markers.
• Non-tissue culture transformation: Direct DNA injection or electroporation techniques reduce reliance on regeneration capacity and are suitable for difficult-to-regenerate crops (such as cassava).

Commercialized GM Crops and Their Global Impact

Overview of Major GM Crops in the Market

The global GM crop market is centered around soybean, corn, cotton, and canola. Data from 2018 showed that GM soybean cultivation accounted for 50% of the global GM crop area, followed by corn (30%), cotton (25%), and canola (20%). China, the United States, Brazil, India, and other countries are major producers. The United States has an average adoption rate of 93% for GM soybean, corn, and cotton, demonstrating the efficiency of technology adoption. Furthermore, the area of biotech crops with stacked traits (such as drought and insect resistance) has grown significantly, accounting for 42% of the global biotech crop area in 2018, demonstrating strong farmer demand for multifunctional traits.

Socioeconomic Benefits and Yield Growth

The commercialization of biotech crops has significantly boosted agricultural productivity. Since their first commercialization in 1996, biotech crop yields have increased 87-fold, providing a stable food supply for 7.4 billion people worldwide. For example, herbicide resistance reduces pesticide use and lowers production costs, while insect resistance reduces pest losses and increases crop survival rates. Furthermore, biotech technology has promoted sustainable agricultural development. For example, salinity-alkaline tolerance has been added to the energy grass switchgrass, enabling its successful cultivation in saline-alkali lands and contributing to ecological restoration.

Challenges and Future Outlook

Technical Bottlenecks and Challenges

Despite significant progress in transgenic technology, its application still faces multiple challenges. First, the bottleneck of transformation efficiency is a core issue. Many crops (such as rice and wheat) are insensitive to traditional tissue culture and regeneration conditions, resulting in transformation rates as low as 10% or below. Second, public concerns arise about off-target effects and the risk of gene flow. For example, gene editing technologies like CRISPR-Cas9 may accidentally insert non-target genes or spread to wild populations through pollen. Furthermore, insufficient public acceptance limits the technology's widespread adoption, especially in areas with religious or cultural sensitivities.

Emerging Trends and Solutions

Future technological development will focus on the following areas:

• De novo domestication and stacked traits: Through genome editing and multi-gene stacking, crops with stacked traits that combine stress tolerance, high yield, and enhanced nutrition can be developed, such as drought-tolerant and high-protein corn.
• Non-tissue culture transformation methods: Direct DNA delivery and optimized Agrobacterium-mediated transformation methods can reduce reliance on complex tissue culture and improve transformation efficiency.
• Automation and high-throughput screening: Combining artificial intelligence and high-throughput sequencing technologies accelerates the screening and verification of transgenic plants, reducing R&D costs.

Global Cooperation and Policy Coordination

To achieve the sustainable development of transgenic technology, international cooperation and regulatory coordination are necessary. For example, establishing unified off-target effect detection standards and gene flow monitoring systems can enhance public trust. At the same time, developing countries need to narrow the technological gap with developed countries through policy support and capacity building.

Conclusion

As a core driver of modern agriculture, plant transformation technology has profoundly changed the paradigm of crop improvement. From early Agrobacterium-mediated transformation to modern gene editing, continuous technological advancements have provided innovative solutions to address food security, climate change, and resource scarcity. However, its development must balance technological innovation with social ethics. International cooperation and policy coordination should be used to promote the sustainable application of transgenic technology.

Plant Transformation Services

At Lifeasible, we specialize in providing cutting-edge plant transformation services to meet your agricultural and biotechnology research needs. Our expertise in genetic engineering and gene editing technologies enables us to provide precise and efficient transformation solutions for a wide range of plant species. Whether you're looking to enhance crop traits such as stress tolerance, nutritional value, or pest and disease resistance, we offer comprehensive services to meet your specific project needs.

Explore the Various Plant Species for Which We Offer Transformation Services

To help you select the right transformation service, we've provided a detailed table listing the plant species we support. This table will help you understand the scope of our capabilities.

Table1. Plant Species Covered by Lifeasible Transformation Services

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Frequently Asked Questions (FAQs)

Q: What is plant transformation technology?

A: Plant transformation technology refers to the introduction of foreign genes into the plant genome, thereby modifying its genetic traits. It is used for crop improvement and biotechnology advancements.

Q: How does Agrobacterium-mediated transformation work?

A: This method uses Agrobacterium tumefaciens to integrate foreign genes into the plant genome via the T-DNA region of the Ti plasmid and is primarily applicable to dicotyledons.

Q: What are stable and transient transformation?

A: Stable transformation involves the integration of foreign genes into the plant genome for heritability, while transient transformation is temporary and suitable for short-term research.

Q: What are the applications of plant transformation in agriculture?

A: Applications include enhancing resistance to biotic and abiotic stresses, improving nutritional value, enhancing herbicide resistance, increasing crop yield, and extending shelf life.

Q: What challenges does plant transformation technology face?

A: Challenges include low transformation efficiency, potential off-target effects, the risk of gene flow, and public acceptance issues.

Q: What are the socioeconomic benefits of commercialized genetically modified crops?

A: Genetically modified crops have increased agricultural productivity, reduced pesticide use, lowered production costs, and promoted sustainable agricultural development.

References

1. Su, W., et al. (2023). Technological development and application of plant genetic transformation. International Journal of Molecular Sciences, 24(13), 10646. DOI: 10.3390/ijms241310646.
2. Gao, C. (2021). Genome engineering for crop improvement and future agriculture. Cell, 184(6), 1621-1635. DOI: 10.1016/j.cell.2021.01.005.
3. Imran, Q. M., et al. (2021). Abiotic stress in plants; stress perception to molecular response and role of biotechnological tools in stress resistance. Agronomy, 11(8), 1579. DOI: 10.3390/agronomy11081579.
4. Koul, B. (2022). Plant Transformation Techniques. In: Cisgenics and Transgenics. Springer, Singapore. DOI: 10.1007/978-981-19-2119-3_1.
5. Quezada, G.D.Á., Ijaz, S., Malik, R. (2024). Genetic Transformation in Plants: Methods and Applications. In: Ijaz, S., Ul Haq, I., Mohamed Ali, H. (eds) Trends in Plant Biotechnology. Springer, Singapore. DOI: 10.1007/978-981-97-0814-7_2.
6. Tsakirpaloglou, N., et al. (2023). Guidelines for performing CRISPR/Cas9 genome editing for gene validation and trait improvement in crops. Plants, 12(20), 3564. DOI: 10.3390/plants12203564.

Soybean Transformation

Rice Transformation

Maize Transformation

Sugarbeet Transformation

Wheat Transformation

Arabidopsis Transformation

Tobacco Transformation

Tomato Transformation