Genome or gene editing includes several new techniques to help scientists precisely modify genome sequences. The techniques also enables us to alter the regulation of gene expression patterns in a pre-determined region and facilitates novel insights into the functional genomics of an organism. Emergence of genome editing has brought considerable excitement especially among agricultural scientists because of its simplicity, precision and power as it offers new opportunities to develop improved crop varieties with clear-cut addition of valuable traits or removal of undesirable traits. Research is underway to improve crop varieties with higher yields, strengthen stress tolerance, disease and pest resistance, decrease input costs, and increase nutritional value. Genome editing encompasses a wide variety of tools using either a site-specific recombinase (SSR) or a site-specific nuclease (SSN) system. Both systems require recognition of a known sequence. The SSN system generates single or double strand DNA breaks and activates endogenous DNA repair pathways. SSR technology, such as Cre/loxP and Flp/FRT mediated systems, are able to knockdown or knock-in genes in the genome of eukaryotes, depending on the orientation of the specific sites (loxP, FLP, etc.) flanking the target site. There are 4 main classes of SSN developed to cleave genomic sequences, mega-nucleases (homing endonuclease), zinc finger nucleases (ZFNs), transcriptional activator-like effector nucleases (TALENs), and the CRISPR/Cas nuclease system (clustered regularly interspaced short palindromic repeat/CRISPR-associated protein). The recombinase mediated genome engineering depends on recombinase (sub-) family and target-site and induces high frequencies of homologous recombination. Improving crops with gene editing provides a range of options: by altering only a few nucleotides from billions found in the genomes of living cells, altering the full allele or by inserting a new gene in a targeted region of the genome. Due to its precision, gene editing is more precise than either conventional crop breeding methods or standard genetic engineering methods. Thus this technology is a very powerful tool that can be used toward securing the world's food supply. In addition to improving the nutritional value of crops, it is the most effective way to produce crops that can resist pests and thrive in tough climates. There are 3 types of modifications produced by genome editing; Type I includes altering a few nucleotides, Type II involves replacing an allele with a pre-existing one and Type III allows for the insertion of new gene(s) in predetermined regions in the genome. Because most genome-editing techniques can leave behind traces of DNA alterations evident in a small number of nucleotides, crops created through gene editing could avoid the stringent regulation procedures commonly associated with GM crop development. For this reason many scientists believe plants improved with the more precise gene editing techniques will be more acceptable to the public than transgenic plants. With genome editing comes the promise of new crops being developed more rapidly with a very low risk of off-target effects. It can be performed in any laboratory with any crop, even those that have complex genomes and are not easily bred using conventional methods.

Genome editing tools have rapidly been adopted by plant scientists for crop improvement. Genome editing using a multiplex sgRNA-CRISPR/Cas9 genome editing system is a useful technique for crop improvement in monocot species. In this study, we utilized precise gene editing techniques to generate wheat 3’(2’), 5’-bisphosphate nucleotidase (TaSal1) mutants using a multiplex sgRNA-CRISPR/Cas9 genome editing system. Five active TaSal1 homologous genes were found in the genome of Giza168 in addition to another apparently inactive gene on chromosome 4A. Three gRNAs were designed and used to target exons 4, 5 and 7 of the five wheat TaSal1 genes. Among the 120 Giza168 transgenic plants, 41 lines exhibited mutations and produced heritable TaSal1 mutations in the M₁ progeny and 5 lines were full 5 gene knock-outs. These mutant plants exhibit a rolled-leaf phenotype in young leaves and bended stems, but there were no significant changes in the internode length and width, leaf morphology, and stem shape. Anatomical and scanning electron microscope studies of the young leaves of mutated TaSal1 lines showed closed stomata, increased stomata width and increase in the size of the bulliform cells. Sal1 mutant seedlings germinated and grew better on media containing polyethylene glycol than wildtype seedlings. Our results indicate that the application of the multiplex sgRNA-CRISPR/Cas9 genome editing is efficient tool for mutating more multiple TaSal1 loci in hexaploid wheat.

Plant defensins are small cysteine-rich peptides which belong to a group of pathogenasis related defense mechanism proteins. The proteins inhibit the growth of a broad range of microbes and are highly stable under extreme environmental stresses. Tomato cultivation is affected by fungal disease such as Fusarium wilt. In order to overcome fungal damages, transgenic tomato plants expressing the Medicago sativa defensin gene MsDef1 under the control of the CaMV 35S promoter were developed. The Fusarium-susceptible tomato (Lycobersicum esculentum Mill) cultivar CastleRock was used for transformation to acquire fungal resistance. Hypocotyl with a part of cotyledon (hypocotyledonary) for young tomato seedlings were used as an explant material and transformation was performed using the biolistic delivery system. Bombarded shoots were selected on regeneration medium supplemented with hygromycin and suitable concentrations of BA, zeatin ripozide and AgNO3. Putative transgenic plantlets of T0 were confirmed by PCR analysis using primers specific for the transgene and the transformation frequency obtained was 52.3%. Transformation and transcription of transgenes were confirmed in T1 by PCR, Southern hybridizations, and reverse-transcription PCR (RT-PCR). The copy numbers of integrated transgene into tomato genome ranged between 1-3 copies. Greenhouse bioassay was performed on the transgenic T1 and T2 young seedlings and non-transgenic controls by challenging with a vigorous isolate of the fungal pathogen Fusarium oxysporum f. sp. Lycopersici. The level of fungal infectivity was determined using RT-PCR with tomatinase specific primers. Transgenic lines were more resistant to infection by fusarium than the control plants. These results indicated that overexpressing defensins in transgenic plants confer resistance to fungal pathogens.

The highly conserved Sal1 encodes a bifunctional enzyme with inositol polyphosphate-1-phosphatase and 3′ (2′), 5′-bisphosphate nucleotidase activity and has been shown to alter abiotic stress tolerance in plants when disrupted. Precise gene editing techniques were used to generate Sal1 mutants in hexaploid bread wheat. The CRISPR (Clustered Regulatory Interspaced Short Palindromic Repeats) Cas9 system with three guide RNAs (gRNAs) was used to inactivate six Sal1 homologous genes within the Bobwhite wheat genome. The resulting mutant wheat plants with all their Sal1 genes disabled had slimmer stems, had a modest reduction in biomass and senesced more slowly in water limiting conditions, but did not exhibit improved yield under drought conditions. Our results show that multiplexed gRNAs enabled effective targeted gene editing of the Sal1 gene family in hexaploid wheat. These Sal1 mutant wheat plants will be a resource for further research studying the function of this gene family in wheat.

The sugarcane (Saccharum X officinarum) is one of the most important crops used to produce sugar and raw material for biofuel in the world. One of the main causes for sucrose content and yield losses is the attack by insect. In this investigation, cry1Ac gene was introduced into sugarcane variety GT54-9(C9) using the Agrobacterium tumefaciens transformation method for transgenic sugarcane production presenting insect-resistance. The A. tumefaciens strain GV1303 including pARTcry1Ac vector was used for the production of transformed sugarcane. The Bacillus thuringiensis cry gene were successfully used to produce transgenic plants used for the improvement of both agronomic efficiency and product quality by acquiring insect resistance. PCR and Southern hybridization techniques were used to confirm the cry1Ac gene incorporation into sugarcane genome. Transformation percentage was 22.2% using PCR analysis with specific primers for cry1Ac and npt-II (Neomycin phosphotransferase) genes. The expression of cry1Ac gene was determined using reverse transcriptase polymerase chain reaction (RT-PCR), QuickStix test, and insect bioassays. Bioassays for transformed sugarcane plants showed high level of toxicity to Sesamia cretica giving 100% mortality of the larvae. Sugarcane insect resistance was improved significantly by using cry1Ac gene transformation.

Promoting sustainable agriculture and improving nutrition are the main United Nation’s sustainable development goals by 2030. New technologies are required to achieve zero hunger, and genome editing technology is the most promising one. In the last decade, genome editing (GE) using the CRISPR/Cas system has attracted researchers as a safer and easy tool for genome editing in several living organisms. GE has revolutionized the field of agriculture by improving biotic and abiotic stresses and yield improvement. GE technologies were developed fast lately to avoid the obstacles that face GM crops. GE technology, depending on site directed nuclease (SDN), is divided into three categories according to the modification methods. Developing transgenic-free edited plants without introducing foreign DNA meet the acceptance and regulatory ratification of several countries. There are several ongoing efforts from different countries that are rapidly expanding to adopt the current technological innovations. This review summarizes the different GE technologies and their application as a way to help in ending hunger.