Genome-editing, a recent technological advancement in the field of life sciences, is one of the great examples of techniques used to explore the understanding of the biological phenomenon. Besides having different site-directed nucleases for genome editing over a decade ago, the CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR-associated protein) based genome editing approach has become a choice of technique due to its simplicity, ease of access, cost, and flexibility. In the present review, several CRISPR/Cas based approaches have been discussed, considering recent advances and challenges to implicate those in the crop improvement programs. Successful examples where CRISPR/Cas approach has been used to improve the biotic and abiotic stress tolerance, and traits related to yield and plant architecture have been discussed. The review highlights the challenges to implement the genome editing in polyploid crop plants like wheat, canola, and sugarcane. Challenges for plants difficult to transform and germline-specific gene expression have been discussed. We have also discussed the notable progress with multi-target editing approaches based on polycistronic tRNA processing, Csy4 endoribonuclease, intron processing, and Drosha ribonuclease. Potential to edit multiple targets simultaneously makes it possible to take up more challenging tasks required to engineer desired crop plants. Similarly, advances like precision gene editing, promoter bashing, and methylome-editing will also be discussed. The present review also provides a catalog of available computational tools and servers facilitating designing of guide-RNA targets, construct designs, and data analysis. The information provided here will be useful for the efficient exploration of technological advances in genome editing field for the crop improvement programs.

Over the past decades, numerous efforts were made towards the improvement of cereal crops mostly employing traditional or molecular breeding approaches. The current scenario made it possible to efficiently explore molecular understanding by targeting different genes to achieve desirable plants. To provide guaranteed food security for the rising world population particularly under vulnerable climatic condition, development of high yielding stress tolerant crops is needed. In this regard, technologies upgradation in the field of genome editing looks promising. Clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 is a rapidly growing genome editing technique being effectively applied in different organisms, that includes both model and crop plants. In recent times CRISPR/Cas9 is being considered as a technology which revolutionized fundamental as well as applied research in plant breeding. Genome editing using CRISPR/Cas9 system has been successfully demonstrated in many cereal crops including rice, wheat, maize, and barley. Availability of whole genome sequence information for number of crops along with the advancement in genome-editing techniques provides several possibilities to achieve desirable traits. In this review, the options available for crop improvement by implementing CRISPR/Cas9 based genome-editing techniques with special emphasis on cereal crops have been summarized. Recent advances providing opportunities to simultaneously edit many target genes were also discussed. The review also addressed recent advancements enabling precise base editing and gene expression modifications. In addition, the article also highlighted limitations such as transformation efficiency, specific promoters and most importantly the ethical and regulatory issues related to commercial release of novel crop varieties developed through genome editing.

Solanum lycopersicum cv. Pusa Ruby (PR) is a superior tomato cultivar routinely used as a model tomato variety. Here, we report a reference-guided genome assembly for PR, covering 97.6% of the total single-copy genes in the solanales order. The PR genome contains 34,075 genes and 423,288 variants, out of which 127,131 are intragenic and 1232 are of high impact. The assembly was packaged according to PanSol guidelines (N50 = 60,396,827) with the largest scaffold measuring 85 megabases. The similarity of the PR genome assembly to Heinz1706, M82, and Fla.8924 was measured and the results suggest PR has the lowest affinity towards the hybrid Fla.8924. We then analyzed the regeneration efficiency of PR in comparison to another variety, Pusa Early Dwarf (PED). PR was found to have a high regeneration rate (45.51%) and therefore, we performed allele mining for genes associated with regeneration and found that only AGAMOUS-LIKE15 has a null mutation. Further, allele mining for fruit quality-related genes was also executed. The PR genome has an Ovate mutation leading to round fruit shape, causing economically undesirable fruit cracking. This genomic data can be potentially used for large scale crop improvement programs as well as functional annotation studies.