**6. Conclusion and future prospects**

*Genetic Engineering - A Glimpse of Techniques and Applications*

such issues associated with the existing delivery system.

plasts (cell-wall free) with the transformation (85%) efficiency.

in the genetic engineering of the plant system.

with animal system [90, 91]. Usually, two modes of transformation of genes exist in plants system: (1) cargo delivery that depends on the delivery techniques and (2) regeneration by using transformed plants that depends on the tissues, optimization of the protocols, and complicated hormone mixtures. However, the existing technologies have a lot of limitations such as less transformation, high toxicity, and DNA integration into host genome. The grand challenges of genes/biomolecules cargo delivery within the plants system due to the presence of rigid and multilayered plant-cell wall, thereby slower transformation of genes/biomolecules within the plants. To overcome such issues, two approaches have been developed and used for transformation of genes/biomolecules within the plants: (1) *Agrobacterium*mediated delivery system and (2) biolistic particle delivery (DNA bombardment). However, these strategies also have various drawbacks/limitations such as species dependence (changing the species changed the transformation efficiency), required regeneration from tissues, thereby time consuming and less efficiency, and *Agrobacterium*-mediated genes/biomolecule transformation might introduce foreign genetic materials. The *Agrobacterium*-mediated genes/biomolecules might cause disruption of genes/poor/unstable gene expression due to the random DNA integration. The DNA integration might be prevented by using nonintegrated viruses or plasmid deficient in transfer DNA insertion [90, 92]. Therefore, these two strategies are more preferred tools in comparison with other conventional methods. In this context, nanotechnology might be an alternative tool to resolve

Various nanomaterial-based plant delivery systems focus on the synthesis of nanomaterials, agrochemical delivery system, micronutrient delivery system, translocation of nanomaterials that augmented the growth of plants by using metalbased nanoparticles, CNTs, CNFs, quantum dots, graphene and its derivatives, and fullerenes. On the other hand, some nanomaterials exhibited phytotoxicity due to the oxidative stress and vascular blockage, damaging the structural DNA. Recently, Demirer et al. [90] developed nanomaterial-mediated biomolecule delivery system for gene expression and silencing of the plant system. For this, grafting of DNA on covalently functionalized pristine SW-CNTs and MW-CNTs was done to produce effective DNA delivery with strong expression of protein in mature *Eruca sativa* (arugula) leaves. The DNA is delivered in plant nucleus with the CNTs and also silencing of functional gene, separately. The grafting of DNA is done on CNTs due to the π-π stacking; the SDS is replaced by adsorption DNA by using the dialysis process. The produced DNA-CNT-based delivery system is comparable to *Agrobacterium*-mediated delivery system. The study also suggested that the produced CNT-based delivery system efficiently expresses protein in arugula proto-

Zhao et al. [93] developed nanoparticle-mediated genetic transformation. For this, they formed the complex of DNA-nanoparticles and delivered into the pollen grains by using magnetic force. The produce approaches to be moderate with insignificant toxicity, genetically stable and transformed plants. These studies suggested that nanomaterial-based delivery system plays a significant role in the advancement

In general, genetic engineering of the plant system is more complicated compared with animal system. The approach of the genes/biomolecule transformation within the plants still remains a concern due to the multi-layer and rigid cell-wall. There is lack of effective delivery of the diverse genes/biomolecules within the plant system without damaging the tissues. The nanotechnology might be an alternative tool in the advancement of the genetic engineering in plant systems that resolve such delivery challenge of genes/biomolecules, thereby increasing the utility of

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genetic engineering.

Polymeric nanocomposites own distinct features of biodegradability and biocompatibility, which makes it an ideal material to be used in crop protection and micronutrient delivery in the agriculture field. The reactive nanomaterials have been used in various applications because of their functional groups and characteristics; therefore, ENMs might have the potential ability to be used in different applications including agriculture. Moreover, encapsulation of polymers with different nanomaterials like metal/metal-oxide and carbon-based nanomaterials enhanced the controlled release behaviors, biocompatibility, and simple use. Therefore, they are efficiently used in various applications mainly in agriculture. Additionally, uptake, accumulation, and translocation ability of the nanomaterials mainly depend on surface charges, size, and chemical nature of the materials. On the one hand, polymeric coating of nanomaterials might change the functionality and surface charge; therefore, polymeric composite might efficiently translocate within the plants. With regard to advancement in the genetic engineering, nanomaterials might be alternative tools that efficiently delivered genes/biomolecules. Therefore, polymeric nanocomposite enhances the utility of genetic engineering in plant system. As discussed in the text, CNFs is the next generation fertilizer that can easily deliver micronutrients and biomolecules within the plant. However, transformation of these research into field, some issues must be discuss or detailed studies required; (1) cost of the nanofertilizers, (2) safety concern like health/environmental toxicity, and (3) easy applications. We need to do more research in such agricultural areas for easy applicability in the field.
