In agroecology, the integrated approach in production of crops and conceptual marketing management draws holistic economic concern. As reported by “Food and Agriculture Organization” (FAO, 2017), the crop failure be falls mainly due to biotic and abiotic factors, significantly influencing the economic values of crops (Gautam and Kumar, 2020). Environmental and climate changes, causing frequent flood, droughts, temperature variations, higher soil salinity, use of harmful chemicals as additives/fertilizers, pathogen triggered diseases, deteriorate the plant health and affects directly to the crop quality and yield (Bing, 2020). Typically, breeders follow customary methods along with marker-assisted selection to introduce new traits in plants. Some chemical compounds and irradiation techniques are also employed to attain desirable traits; however, they often lead to random mutations in crop genomes.

The customary methods have certain drawbacks such as non-specificity and the generation of mutations with abundance of nucleotides (Mao et al., 2019). Natural chemical compounds from plant source that are essential to the pharmaceutical industry can only be acquired in limited quantity from normal plants compared to genomically edited plants. Moreover, the rational methods are time-consuming; consequently, breeders are unable to grow plants with desired trait in deadline. Supplying sufficient food and other plant-based chemical constituents to ever-growing population is challenging; and quite perplexing topic in near future. The CRISPR/Cas9 genome editing (GE) with nanotechnology-based protocols can potentially challenge these obstacles.

The targeted genomic engineering can be extremely beneficial to agriculture. If the function of a specific gene is known, it can be over expressed or suppressed to obtain the desired trait. Eventually, the crops that are capable of withstanding biotic and abiotic stresses can be easily developed using CRISPR/Ca9 and nanotechnology. Similarly, undesired genes can also be silenced by using these technologies, which would allow expression of only the desirable traits to eventually obtain higher yield. It will be an extremely difficult task to improve crop refinement without genomic engineering. Conventional breeding programs would require longer duration to introduce new quality-related traits or disease-tolerance traits. Therefore, genomic engineering is a striking technology for the future development of agricultural crops with nutrition. Nanotechnology-based protocols has new sets of advanced applications in agriculture and biomedicine; typically, the nano-sized particles are used to deliver the task obtain desirable results in crop development. Coalescing biotechnology and nanotechnology approaches, including GE, have more benefits than customary breeding to improve the development of food crops, which can naturally overcome biotic and abiotic stress along with enhancing the yield. This brief review mainly focuses the importance of genomic engineering in agriculture, and the progress in developing mutant plants using sequence-specific nucleases (SSNs). Furthermore, the procreation of genomically edited crops are already developed in agricultural biotechnology, the main objective, limitations and prospective challenges of GE are highlighted including the role of nanotechnology-based methods in crop refinement.

Sequence specific nucleases are mainly used for precise gene editing in plants and animals. It can create mutations at desired loci in multiple genes via addition, deletion, and alternation of sequences (Songstad et al., 2017). To generate the mutant plants, firstly SSNs requires to be articulated in cells; subsequently, recognizing a specific DNA sequence to make the double stranded break. We can classify the SSNs into four major classes, namely (i) CRISPR-Cas9, (ii) Zinc finger nucleases (ZFN), (iii) Meganucleases, and (iv) Transcription activator-like effector nucleases (TALENs). These methods can be effectively utilized for the GE technique. CRISPR-Cas9 is derived from the adaptive immune systems of bacteria; in this mechanism, abounding components come into play to perform the GE. Zinc finger nucleases (ZFNs) are the enzymes that have been characterized 77 candidate two-finger modules (Urnov et al., 2005; Miller et al., 2007). Meganucleases are the microbial enzymes (Smith et al., 2006) that can 76 distinguish and more than 14 nucleotides for cleavage. TALENs have been developed by combining the FokI nuclease domain with TALE proteins of Xanthomonas (Christian et al., 2010). CRISPR/Cas9 is one among the four classes of SSNs that can be used for GE (Songstad et al., 2017). GE technologies by adopting CRISPR/Cas9 methodology was investigated in 1987 and its functional application in human cell was reconnoitered in 2013 (Mali et al., 2013). Hsu et al. (2014) discussed the detailed challenges and its future prospective. Consequently, the GE technology was successfully implemented in crop refinement of soybean plant (Cai et al., 2015). Once the SSNs construct is incorporated into the plant genome, they are expressed at a distal site, while the remaining construct is removed by crossing the plant to obtain a mutated plant with no transgene. For GE with the CRISPR/Cas9 system, it is essential to deliver sgRNA and Cas9 proteins into the target cells. Expression vectors or microinjected RNA/mRNA (for Cas9) are usually used to express the sgRNA and Cas9 protein in the plant cells. The CRISPR/CAS9 technology has been performed in plant cells by using electroporation, Agrobacterium-mediated transformation, shotgun methodologies, and polyethylene glycol-mediated routes (Jiang et al., 2013; Li et al., 2013; Mao et al., 2013; Nekrasov et al., 2013; Shan et al., 2013). The double stranded break can be fixed either by homology directed repair (HDR) or non-homologous end joining (NHEJ) (Hsu et al., 2014). Recently, RNA viruses have been used to deliver hairpin RNAs for gene silencing, which is another reported technique to incorporate SSNs into the plant cells (Lacomme, 2015). Before the integration into plant’s genomic DNA, SSNs are transiently expressed from viral vectors into mRNAs and its respective proteins. The ability to modify the genes to modulate specific traits and homologous recombination (HR) allows plant to metabolize in a manner that develops their resistance to biotic and abiotic stresses. Plants synthesized by fast-growing genome engineering, generally exhibit higher crop yields because of its higher ability for photosynthesis. Precise altering the DNA sequence is extremely important to comprehend the achievable challenges insynthetic biology (Abdallah et al., 2014; Carroll, 2014).

Transgenesis profoundly involving genetic addition without changing the genetic pool to create specific traits for agricultural/agroeconomical benefits. In the present scenario, providing food security to a mounting populace is one of the major challenges in this modern world. Thenceforth, the food fructification needs to be increased over 80–90%. Furthermore, the food production is declining every year due to extreme weather conditions, climate change, global warming, farmland availability, and cumulative biotic and abiotic problems. To overcome these challenging obstacles, gene modification technique in food crops will be the superlative method to achieve the targeted tasks (FAO, 2017). The gene knockout step is a critical stage and it directly influence the phenotype. Wada et al. (2020) demonstrated genomic manipulation without the introduction of DSBs. In this modular approach, a dead Cas9 variant (dCas9) binds to the target sequence; however, it does not cleave the double-stranded DNA (Qi et al., 2013; Wang et al., 2016a; Adli, 2018). Generating gene knockouts using SSNs facilitates genetic analyses and the study of important gene functions, which will eventually help with crop improvement. The first mutation generated using SSNs were of the IPK1 gene in maize, which is catalyzed in the last step during phytate biosynthesis (Liang et al., 2014). Knocking out this gene helps in removing unwanted metabolites; eventually contributing to the accumulation of valuable biosynthetic intermediates. Prime editing is another significant approach for genomic manipulation that has been indicated in mammalian and yeast cells (Anzalone et al., 2019). Certain crops such as grains, vegetables, and fruits are vitally important to maintain global food securities and sustainable system.

Homologous recombination is a challenging process wherein chromosome can chasm by the nuclease that must be coordinated with the distribution of the DNA repair template. Initially, HR in plants was confirmed by the incorporation of marker genes at detailed chromosomal sites. Targeted transgene insertion into the euchromatin should provide promising results in plants with high transgene expression; therefore, it is a great improvement over the integration by the traditional Transgenesis (Neelendra et al., 2018). Moreover, the insertion of multiple genes at the same site will facilitate their transfer to a single mendelian locus, when the plant is crossed. Further efforts are required to familiarize plentiful genes into the germplasm by breeding. Targeted gene inclusion through HR using different SSNs has been verified in tobacco, maize, and rice (Abdallah et al., 2014; Wang et al., 2016b). In order to develop new plant varieties, some techniques have been used that are controlled by process-based regulatory frameworks. Most of the farmers, globally aim to cultivate crops that are tolerant to drought, high salinity, and diseases, with appreciable yield. Therefore, the researchers in agriculture domain are actively looking for transgenic technology and CRISPR/CAS9 to achieve targeted tasks. Guidelines and process-based regulations have been formulated and implemented by the United Nations Food and Agricultural Organization adopted by European and South American countries (FAO, 2017).

Genomic engineering has been encountered broad range of applications to introduce targeted alterations of the plant’s genome to acquire desired function (Hsu et al., 2014). The NmeCas9 (Neisseria meningitidis) recognizes an 8-mer PAM (5′-NNNNGATT) sequence hence it can progress the target particularity, whereas SaCas9 recognizes a 6-mer PAM sequence (5′-NNGRRT) (Xing et al., 2014; Ma et al., 2015). This technique involves introducing mutations, and harnessing transgene supplementation for gene therapy and livestock improvement. CMV (Califlower mosaic Virus), AtUBO (Arabidopsis UBO) OsUBO (Oriza sativa UBO), LTR (Long terminal Repeat), OsnoRNA U3 (Oriza sativa snoRNA U3), AtU6 (Arabidopsis U6), OsUBI (Oriza sativa Ubiquitin), ZmUBI (Zea mays UBI), cauliflower mosaic virus 35S promoters have been used to promote Cas9 expression in plants and, more than 30 empty gRNA backbones in binary vectors was supplied by Addgene (Belhaj et al., 2013; Feng et al., 2013; Mao et al., 2013; Nekrasov et al., 2013; Upadhyay et al., 2013; Xie and Yang, 2013). Tissue specific promoters can also be used in CRISPR/Cas9 technology to edit genomes; for instance, Wang et al. (2015) used the promoter of the egg cell-specific EC1.2 gene, to initiative of Cas9. The germ line-specific SPOROCYTELESS and embryo-specific promotor DD45 (Mao et al., 2016). The promotor AtDMC1 involved in meiotic recombination (Xu et al., 2018). The pLAT52-GT for Pollen tissues (Mao et al., 2016), and pDD45-GT for egg cell-early embryo tissues (Mao et al., 2016), INCURVATA2 for dividing tissue-targeted site-directed mutagenesis (Hyun et al., 2015), and the YAO promoter for cell-division specific tissues (Yan et al., 2015). In order to knock out the expressions of multiple genes cassettes can be inserted into one plasmid, thereby guiding the Cas9 to different targets (Xing et al., 2014; Ma et al., 2015; Oz et al., 2021). Although this GE technology has key advantages, there are some negative shades involving ethical issues concerning to the disruption of ecological balance (Tavakoli et al., 2021). To overcome this, different methods including nanotechnology-based methods are being implemented to insert or silence the genes in plant cells (Nadakuduti and Enciso-Rodríguez, 2021).

Despite animal-based food consumption, the global ecosystem mainly be contingent to the agro-based crops including herbivorous animals. In concern to this, extensive research findings are mainly focusing on developing nutritional cereal/vegetables/fruits/nuts (FAO, 2017). Cultivating nutritional crops mainly depends on the nature and the fertility of the soil. Sources of soil nutrients are not same and is mainly depending on the presence of organic matter. In order to overcome this nutrient deficiency, CRISPR/Cas9 technology is extremely useful to grow cereal/vegetables/fruits/nuts with high nutritional values. Recently, genomic engineering utilizes TALENs, Cas9, dCas9, and Cre inserting into the cells. The use of these proteins in human cell lines have also been verified in vitro and in vivo (Zuris et al., 2015). HR (Sun et al., 2016) mediates the modifications made by the engineered nucleases. To date, plentiful of food-based crops have been modified to obtain good nutritional values in vegetables and fruits (Kathleen, 2015). Varieties of fruits comprised of different nutrients and biologically active compounds that are necessary in daily healthy diet. Recently, Dalla et al. (2019), Wan et al. (2021), and Xu et al. (2021) have discussed the importance of gene editing in fruits and vegetables to get nutritional rich cereal/vegetables/fruits/nuts, which are beneficial to maintain good health.

Genome-wide association studies (GWAS) have been employed to identify specific locations in the genome that can anchorage polygenic diseases such as Alzheimer’s, diabetes, autism, and schizophrenia. This technique is crucial in biomedical field to treat various diseases, including the removal of HIV genome (Ebina et al., 2013; Liao et al., 2015; Kaminski et al., 2017). The ex vivo and in vivo GE of neurons, immune cells, and endothelial cells has been successfully reported in mice (Platt et al., 2014), which are challenging to modify the sensitive cells and its effective editing. Researchers have introduced resistance against malaria by editing the DNA in Anopheles mosquitoes (Gantz et al., 2015; Hammond et al., 2016; Macias et al., 2020). Cancer therapy strategies have also been conducted in biomedical field to treat cancerous cell lines (Yao et al., 2015; Khan et al., 2016; Yang H. et al., 2019). Furthermore, AIDS research is still ongoing to engraft Cas9-modified CCR5-human hematopoietic stem cells and progenitor cells (Mandal et al., 2014). GE techniques are gaining its impact in promising therapeutics in regenerative medicine. The primary route for disease treatment is direct GE in tissues; some reports have documents the correction of monogenic recessive genetic disorders, such as Duchenne muscular dystrophy (Ousterout et al., 2015), hemophilia (Park et al., 2016) cystic fibrosis (Schwank et al., 2013), and sickle cell anemia (Sun and Zhao, 2014). The Cas9 system has exhibited its efficacy in therapies through the insertion of SSNs into microbial populations using phages and conjugative plasmids (Citorik et al., 2014).

The comprehensive study on rice (Shan et al., 2013), sorghum (Jiang et al., 2013), tobacco (Li et al., 2013), wheat (Wang et al., 2014), tomato (Brooks et al., 2014), maize (Liang et al., 2014), sweet orange (Jia and Wang, 2014), and Arabidopsis (Li et al., 2013), stretches the vast knowledge of specific gene handling and editing. Before SSNs, RNAi technology was used to study the gene function by knocking down the targeted genes, which was not as advantageous as SSNs. Certain characteristics discussed below encompasses the examples for certain horticultural and ornamental plants using the CRISPR system. The color and weight/size ratio of tomato fruit could be developed by the editing of PL and TBG4 genes (Wang et al., 2019a). The SlNPR1 and SlCBF1 genes corresponding to drought and cold tolerance can also be modified (Li et al., 2018b; Li R. et al., 2019) and the fruit ripening transcription factor RIN (Ripening Inhibitor) could be edited, so that the tomato with desired characteristics will be maintained. Albino phenotype and flowering characters could be modified in cabbage using FRI and PDS gene editing (Murovec et al., 2018). In addition, the biosynthesis of Carotenoid pigment can be enhanced in wild cabbage with BoaCRTISO (Carotenoid isomerase) gene editing (Sun B. et al., 2020). By editing DcF3H and DcPDS, DcMYB113 genes, the accumulation of acylated anthocyanins can be enriched in the roots of carrot to afford pigmented purple carrots (Chodacka et al., 2018; Xu et al., 2019). Cucumber mosaic virus (CMV-Z1) and Zucchini yellow mosaic virus (ZYMV) are two major rapidly affecting pathogens, which can severely damage the crop. To overcome this, the pathogenic resistance can be developed/enhanced by editing elF4EF gene (Chandrasekaran et al., 2016). Drought tolerance in an important commercial crop, hot pepper (Capsicum annuum L. syn. chilli) was developed by editing NAC72 gene (Joshi, 2019). Furthermore, to enhance the flower longevity of attractive petunia flowers can be edited using PhACO1, 3, and 4 gene (Xu et al., 2020). The color and the Carotenoid accumulation of Japanese morning glory flower can be edited by altering its related gene InCCD4 (Carotenoid Cleavage Dioxygenase) (Watanabe et al., 2018). Compatibly, the flavonoid biosynthesis could be enriched in Wishbone flower by editing F3H (Flavanone 3- hydroxylase) gene (Nishihara et al., 2018). Moreover, the AhFAD2A and AhFAD2B genes encoding fatty acid desaturases in groundnut have been reported to be edited (Yuan et al., 2019). The TYLCV-IR (Intergenic regions) gene has been modified to overcome the multiple viral diseases in Nicotiana benthamiana. After gene modification plant exhibited resistance to geminiviridaevirus, begomovirus, curtovirus, becurtovirus, eragrovirus, Turncurtovirus, and Topocuvirus (Ali et al., 2015a,b). The resistance against Phytophthora tropicalis in cacao has been overcome by altering the TcNPR3 gene (Fister et al., 2018). Giovannini et al., 2021 have identified the desirable phenotypic characters in ornamental flowers, including flowering induction, floral meristem initiation and organ development, as well as color, fragrance, and shelf life. Flower longevity has been induced in petunia by altering a group of Petunia hybrid 1-aminocyclopropane-1-carboxylate oxidase (PhACO, PhACO1, PhACO3, and PhACO4) genes (Xu et al., 2020). Canker and huanglongbing diseases are the major factors in reducing the productivity of citrus plants; this problem was overcome by modifying the CsLOB1, CsWRKY22, and DMR6 genes by CRISPR/Cas9 system (Peng et al., 2017; Zhang et al., 2018c; Wang et al., 2019b). Selectable marker gene (SMG) systems are critical and play a major role in the identification of transgenic crops. Nowadays, the scientists are considering the SMGs that can affect human and animal health. The gene transferred plants (GMP: Genetically Modified Plants) usually contain the antibiotic resistant gene, so that GM plants should survive and regenerate under antibiotic medium. Whereas, the non-gene transformed plants will not rejuvenate, eventually die under toxic proximity. Although antibiotics have positive health and life prospective in human/animal health, the negative impacts of antibiotic associated diarrhea and pseudomembranous colitis will proliferate the possibilities of subsequent diseases. By consuming those GMP (Fruits and vegetables) for prolonged usage can severely affect the human/animal health (Breyer et al., 2014). Thus, it is imperative to eliminate SMGs from transgenic crops by using CRISPR technology (Yau and Stewart, 2013). Arndell et al. (2019) edited the biosynthesis of 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) for the functional confirmation of EPSPS gene in wheat using CRISPR/Cas9. The soybean storage protein genes were also been edited successfully to observe the efficacy of CRISPR/Cas9 technique using Agrobacterium rhizogenes-mediated hairy root transformation method (Li C. et al., 2019). Acetolactate synthase (ALS) participates in amino acid biosynthesis; this amino acid is targeted by numerous herbicides. These two plant enzymes(EPSPS, ALS, ACCase), and BFP genes, confer herbicide tolerance in plants (Voytas and Gao, 2014; Sauer et al., 2016; Li et al., 2018e). Hybrid paddy are susceptible to bentazon and sulfonylureas and the BEL gene has been mutated using radiation. In the production of hybrid rice, these mutants can be used to prevent contamination in hybrid seed lots (Cantos et al., 2014). Here, the BEL gene was edited using CRISPR-Cas9 technology, and transformed into rice through A. tumefaciens (Xu et al., 2014). Nutrient values have also been increased in vegetables and fruits by knocking out genes using the CRISPR-Cas9 system. Visually attractive flowers possess pleasant aroma because of the presence of anthocyanin, whose expression is regulated by the MYB-bHLH-WD (MBW) complex (Albert et al., 2014; Lloyd et al., 2017). Gibberellin (GA) determines plant height and strigolactone (SL) affects branching of the shoot branching, both of which can be modulated by modifying the biosynthesis or signal transduction of GA and/or SL (D’Halluin et al., 2008).

Unwanted metabolites usually have negative impacts on the crop yield and its quality; the accumulation of these undesired metabolites can be avoided by using GE. Cyanide intoxication, ataxia or partial paralysis, and goiters are caused by cyanide, which is present in cassava (Padmaja, 1995). Glucosinolates, which produced by mustard and cabbage, also possess a high toxic content, were edited (Hannoufa et al., 2014). FAD2 and FAD3 genes produce high oleic acid and low linolenic acid in soybean; however, soybean oil allows the accumulation of monounsaturated fats and reduces the linolenic acid in the seeds (Pham et al., 2012). AtPDS3, AtFLS2, AtADH, AtFT, AtSPL4, and AtBRI1 genes are targeted in Arabidopsis with mutation rates (MRs) from 1.1 to as high as 84.8% in the first generation (Li et al., 2013). The OsPDS and OsBADH2 genes have been knocked out with MRs of 9.4 and 7.1% (Hinge et al., 2015). The DsRED2, DD20, and DD43 genes have been targeted in sorghum with MRs of 33, 59, and 76%, respectively. Similarly, the ZmIPK (13.1%), LIG1, MS26, MS45, and ALS1 genes have been edited in maize with MRs lower than 5% (Liang et al., 2014; Svitashev et al., 2015). TaMLO-A, TaMLO-B, and TaMLO-Dare three homeo alleles that confer powdery mildew resistance and have been edited with the same moderate MR of 5.6% (Wang et al., 2014). In BRI1, JAZ1, and GAI genes mutation frequencies of 26–84% have been observed (Feng et al., 2013). NtPDS and NtPDR6 have been mutated with MRs of 81.8 and 87.5%, respectively (Gao et al., 2015). The squamosa promoter binding protein-like 4 and Flowering Locus T (FT) have been mutated with an MR of 90%, which caused it to exhibit late flowering (Hyun et al., 2015).

Ma et al. (2015), have reported genome modifications at 46 target sites with an average of 85.4% mutations in monocot and dicot plants using either golden gate ligation or Gibson assembly. Using the sgRNA single, double and triple mutants have also been generated for CDKA2, CDKB1, and CDKB2 in rice (Endo et al., 2015). Dort et al. (2020) discussed forest pathosystems; some disease problems were solved using the CRISPR/Cas9 system. Zhang et al. (2014), have also reported mutations in young seedling albino (OsYSA) and OsROC5 genes with MRs of 65–66.7%. Similarly, Wang et al. (2016b), have edited the OsERF922 gene that encodes ERF transcription factors to develop resistance to rice blast disease. Transgenic poplar plants have been modified and phenotypically results revealed an MR of 51% (Fan et al., 2015). El-Mounadi et al. (2020) explained biosafety of genomically edited plants and the applications of CRISPR/CAS9 technology to enhance yield, quality, and nutritional values. The crops and seeds developed using CRISPR/Cas9 technology for cereals, vegetables, ornamental, and fruits plants are shown in Tables 1, 2 and Supplementary Tables S1, S2. Some multinational companies (DuPont, Monsanto, and BASF) had obtained licenses to develop new crops using CRISPR technology (Khurana and Rajarshi kumar, 2019). The sequential steps for CRISPR/Cas9 genetic transformation in plants was sketched in Figure 2 (Manghwar et al., 2019).

Nanotechnology plays a prominent role in biological, medicinal, and pharmaceuticals including plant science. Increasing resilience to biotic and abiotic stress and improving the yield/quality of the crops via gene editing, nanotechnology shares its connotation with CRISPR/Cas9. Nano-fertilizers (Subramanian et al., 2015; Thul and Saragani, 2015) are used in horticultural plants, including vegetables and fruits, and implemented in food crops to enhance the growth, germination rate, and genetic manipulations (Lee et al., 2010; Sheykhbaglou et al., 2010; Dimkpa et al., 2012; Shang et al., 2019; Mittal et al., 2020; Aqsa et al., 2021; Rana et al., 2021). Similarly, plant growth, nutrient uptake from roots, flowering; have also been developed by metal and carbon based nanoparticles (Aqsa et al., 2021). Photosynthesis is an energy conversion process in plants, transforming light energy into chemical energy; however, it does not occur effectively under cloudy conditions and in sun-drenched plants during the rainy season. Consequently, cell mechanisms possibly down regulated. The gold nanoparticles could be beneficial to enhance the light-harvesting capacity, thereby promoting highly excited electron transfer in the chloroplast (So et al., 2015). Environmental factors (abiotic stress) cause biochemical and physiological changes in plants and these are more susceptible to stress. Even in stressful conditions, the use of metallic nanoparticles can increase the anti-oxidative enzyme levels in plants (Mittal et al., 2020; Zhao et al., 2020; Wu and Li, 2021) and reduce the reactive oxygen species (ROS) levels in the mitochondria and chloroplasts to protect the plant (Sun L. et al., 2020; Sun et al., 2021). However, the use of these nanoparticle fertilizers in crop fields not only increase the soil fertility, but also greatly influence the water resource contamination (Naderi and Abedi, 2012; Mittal et al., 2020). Fertilizers containing microorganisms are labeled as biofertilizers, which can activate the plant system and improve the nutrient uptake from soil (Manikandan and Subramanian, 2016). Nano-fertilizers have the similar benefits like biofertilizers (Elias et al., 2019). Moreover, metallic nanoparticles have anti-pathogenic, antifungal, and antibacterial properties (Kah and Hofmann, 2014; Servin et al., 2015), so that they can survive under pathogenic bout under the soil. Brief explanation of using metallic nanoparticles in farming and its benefit in sustainable agriculture is explained in Figure 3 (Mittal et al., 2020).

Nanoparticles acts as a carrier to deliver the necessary materials into plant cells, animal cells, and specific organs for cancer therapy, genetic disease treatment, and to obtain desirable traits in plants (Peng et al., 2019). Lv et al. (2020) and Ahmar et al. (2021) explained the method in which the gene transformation is performed using nanoparticles. Various types of nanoparticles have been used to deliver genetic material into the plant cells through different platforms. These reports have also detailed the merits and demerits of utilizing nanoparticles in gene transfer methods. Typically, the mesoporous silica nanoparticles, carbon nanotubes, gold, and magnetic nanoparticles have been used to deliver plasmid DNA, double-stranded RNA, and siRNA into plant protoplasts or other intact cell lines (Al-Babili and Bouwmeester, 2015; Yao et al., 2016; Peng et al., 2019; Huan et al., 2020; Tsveta et al., 2021). However, Lv et al. (2020) have demonstrated the gene silencing and gene editing in plants with the use of nanoparticles; magnetic nanoparticle-based pollen transformation was used to achieve the task. In this approach, the vector-magnetic nanoparticle complex has been associated with the pollen that is dropped onto the stigma of desired plant flowers. Finally, plants produce the desired seeds by transferring the vector-magnetic nanoparticle complex into the flower stigma. Later, these flowers are modified into fruits, and the seeds were screened on antibiotic plates. The speed breeding protocol was used to obtain T0, T1, and T2 generations of transgenic plants; this breeding program is inexpensive for editing plant genomes and is employed for various Brassica species (Ahmar et al., 2021). Similarly, dsRNA has been loaded into the layered double hydroxide (LDH) clay nanosheets that are non-degradable, non-toxic, and resistant to easy wash. Furthermore, when these nanoparticle-dsRNA complexes are sprayed onto the plant leaves, they immediately attach to the leaf surface and are absorbed by plant viruses to induce RNAi, eventually degrading the targeted plant pathogens or endogenous mRNA can be minimized/eliminated (Lv et al., 2020). Similarly, gene editing has also been demonstrated with small NPs-CRISPR/Cas9 vector complex that was microinjected into the leaves or any other plant parts, which can be proliferated further by tissue culture or other ease protocols (Lv et al., 2020; Duan et al., 2021). Carbon dots-siRNA complex has been used to silence the GFP in tobacco and tomato plants (Schwartz et al., 2020). Demirer et al. (2021) recently demonstrated genome editing in plants using the CRISPR/Cas9 system along with nanoparticles and explained the regeneration, and phenotypic/metabolic changes of genomically edited crops. The methods in which gene expression, silencing, editing, and other applications involving nanoparticles can account for the crop refinement and are briefly explained in the Figure 4 (Peng et al., 2019).

The CRISPR/Cas9 and nanotechnology aspects were successfully implemented so that relevant solicitations in plant biotechnology, food industry, livestock improvement can be achieved (Islam et al., 2020). Biofuel production (Javed et al., 2019), and rectifications of genetic and metabolic diseases can be done by controlling bacterial and viral diseases (Robert et al., 2017; Zafar et al., 2020), controlling bacterial and viral diseases (Zafar et al., 2020).

CRISPR/Cas9 technology is among the best and most cost-effective methods for genomic editing in plants. However, some of its limitations are mentioned below.

Scientists and plant breeding researchers are working to develop high yielding and biotic/abiotic stress resistant variety. CRISPR/Cas9 GE in consolidation with nanotechnology have emerged as an important platform to improve the quality and desirable quality of agricultural crops with appreciable yield. These technologies are significantly susceptible and open new prospects in plant genetics. The nutritional values and quality of health prospects of plants and human could be enriched with broad spectrum of applications including in biomedical domain. The imminent fertility and diminishing useful prokaryotic microorganism issues in soil are delicate issues in addition to biotic problems. Relapsing the aforesaid issues could be feasible using CRISPR/Cas9 GE and nanotechnology aspects, it will be challenging and greatly influence to the crop refinement. Skillful editing of genetic sequence of plant genome could progressively improve the agronomic traits, photosynthetic capacity, and nutritional values. Furthermore, biotic and abiotic stress-induced issues in plants could be configured and minimized. The methodologies can also be diversified to characterize the individual gene functions, thereby improving the genomic sequences of agricultural crops to produce exceptional yield. These findings could point new strategies in agroecology facilitating the sustainable production of nutritional quality food to satisfy the increasing demand of ever-growing population.

CRISPR/Cas9 and nanoparticle complex system is an advanced innovative technique in agricultural crop development/refinement. Thus far, CRISPR system in connotation with nanotechnology has been used to improve the quality and yield of many valuable crops for future benefits. Using nanoparticle based fertilizers/additives; extinct nutrients could be regained, ensuring that the soil nutrients could be maintained. The green, non-toxic nanoparticles is a sagacious approach to increase micro and macronutrient levels in the soil for healthy growth of crops. Chemically derived nanoparticle usage should be eliminated; these can harm not only the crops but also to the environment. Zinc oxide-based nanocomposites and nano-fertilizers have shown appreciable results in crop growth; maintaining the soil salinity and fertility with robust yield. The green protocols to develop Zinc oxide-based green nanocomposites draws the attention of agro-scientific community and it should be given prime importance, so that toxic free effective nano-priming techniques can reduce the soil contamination and improves the seed quality. Metal organic frameworks (MOF) have been recently investigated as delivery systems for CRISPR/Cas9. Non-toxic and eco-friendly MOF 3-dimentional structures with biopolymer conjugates are potentially promising. Furthermore, CRISPR/Cas9-associated nanoparticle complex has been successfully utilized for transformation, silencing, and modification of genes to overcome the existing and expected critical biotic and abiotic issues, thereby producing nutritional-rich crops. This technology can be used to alter the metabolic pathways in plants to obtain desired high-quality secondary metabolites for future usage. Recently, revolutionary changes in this crop refinement program witnessed auspicious results. Nod factors have been shown to increase nitrogen efficiency in legume crops using CRISPR/Cas9 system with nano-technological contrivance. Modifying the Nod signaling pathway in cereal crops should eliminates the use of toxic inorganic fertilizers. The transformation efficiency by Agrobacterium tumefaciens is quite low in some specified plant tissues. Hence, an alternative bacterial system to gene transfer is necessary for ease genome editing in all crops by adopting CRISPR/Cas9-Nanotechnology system. Furthermore, the development of tissue culture free delivery protocols involving direct genomic editing in germplasms and meristematic cells of plants, can yield propitious results. To date, numerous crops have been developed with this combined CRISPR/Cas9-Nanoparticle complex system. It is advisable to framework on the products already developed by this method and ensure their recurrent use in agricultural locales. Extensive studies are recommended to elucidate the complete interactions (plant cell mechanisms) of nanoparticles/nanocomposites in all types of crops. We strongly believe that the products developed by conjoining these technologies will beneficially assist the agro-based researchers to bloom their ideas for innovative crop development/refinement.

BN planned the manuscript outline, wrote the draft, and prepared the tables. GS contributed in writing, reshaping/editing the manuscript and modifying the tables. GS, S-CK, MM, CS, RP, and MK proofread the manuscript. S-YK acquired the funding. S-YK and S-HL supervised the study and revised the manuscript. All the authors reviewed and approved the final version of the manuscript.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.