A prime example of precisely delivered DNA.

Incorporating large fragments of DNA into specific genome positions is an inefficient process even when using the most cutting-edge genome-editing tools. Sun et al. recently described the prime editing-mediated recombination of opportune targets (PrimeRoot) method, which precisely and efficiently integrates large fragments of DNA into plant genomes and has enormous potential in research and agriculture.

Deciphering the differential expression patterns of yield-related negative regulators in hexaploid wheat cultivars and hybrids at different growth stages.

BACKGROUND: Hexaploid bread wheat underwent a series of polyploidization events through interspecific hybridizations that conferred adaptive plasticity and resulted in duplication and neofunctionalization of major agronomic genes. The genetic architecture of polyploid wheat not only confers adaptive plasticity but also offers huge genetic diversity. However, the contribution of different gene copies (homeologs) encoded from different subgenomes (A, B, D) at different growth stages remained unexplored. METHODS: In this study, hybrid of elite cultivars of wheat were developed via reciprocal crosses (cytoplasm swapping) and phenotypically evaluated. We assessed differential expression profiles of yield-related negative regulators in these cultivars and their F1 hybrids and identified various cis-regulatory signatures by employing bioinformatics tools. Furthermore, the preferential expression patterns of the syntenic triads encoded from A, B, and D subgenomes were assessed to decipher their functional redundancy at six different growth stages. RESULTS: Hybrid progenies showed better heterosis such as up to 17% increase in the average number of grains and up to 50% increase in average thousand grains weight as compared to mid-parents. Based on the expression profiling, our results indicated significant dynamic transcriptional expression patterns, portraying the different homeolog-dominance at the same stage in the different cultivars and their hybrids. Albeit belonging to same syntenic triads, a dynamic trend was observed in the regulatory signatures of these genes that might be influencing their expression profiles. CONCLUSION: These findings can substantially contribute and provide insights for the selective introduction of better cultivars into traditional and hybrid breeding programs which can be harnessed for the improvement of future wheat.

ZmGLK36 transcription factor bestows viral resistance in rice and wheat.

Maize rough dwarf disease (MRDD) threatens the sustainable production of major cereal crops. Recently, Xu et al. reported a new resistance gene, ZmGLK36, which promotes MRDD resistance in maize by increasing jasmonic acid (JA)-mediated defence. This discovery provides opportunities to develop resistance to rice black-streaked dwarf virus (RBSDV) in other cereal crops such as rice and wheat.

Methyl-salicylate (MeSA)-mediated airborne defence.

Stressed plants emit a variety of chemicals into the environment, leading to increased pest resistance in neighbouring plants but the genetic and molecular mechanisms of the emissions remain obscure. Recently, Gong et al. identified novel methyl salicylate (MeSA)-mediated airborne defence that confers resistance to neighbouring plants against aphids and viruses.

Salicylic acid-driven innate antiviral immunity in plants.

Pathogenic viruses are a constant threat to all organisms, including plants. However, in plants, a small group of cells (stem cells) protect themselves from viral invasion. Recently, Incarbone et al. uncovered a novel salicylic acid (SA) and RNAi mechanism of stem cell resistance, broadening our understanding of RNAi-mediated antiviral plant immunity.

Dominance of Cotton leaf curl Multan virus-Rajasthan strain associated with third epidemic of cotton leaf curl disease in Pakistan.

Cotton (Gossypium hirsutum) is an economically potent crop in many countries including Pakistan, India, and China. For the last three decades, cotton production is under the constant stress of cotton leaf curl disease (CLCuD) caused by begomoviruses/satellites complex that is transmitted through the insect pest, whitefly (Bemisia tabaci). In 2018, we identified a highly recombinant strain; Cotton leaf curl Multan virus-Rajasthan (CLCuMuV-Raj), associated with the Cotton leaf curl Multan betasatellite-Vehari (CLCuMuBVeh). This strain is dominant in cotton-growing hub areas of central Punjab, Pakistan, causing the third epidemic of CLCuD. In the present study, we have explored the CLCuD diversity from central to southern districts of Punjab (Faisalabad, Lodhran, Bahawalpur, Rahimyar Khan) and the major cotton-growing region of Sindh (Tandojam), Pakistan for 2 years (2020-2021). Interestingly, we found same virus (CLCuMuV-Raj) and associated betasatellite (CLCuMuBVeh) strain that was previously reported with the third epidemic in the central Punjab region. Furthermore, we found minor mutations in two genes of CLCuMuV-Raj C4 and C1 in 2020 and 2021 respectively as compared to its isolates in 2018, which exhibited virus evolution. Surprisingly, we did not find these mutations in CLCuMuV-Raj isolates identified from Sindh province. The findings of the current study represent the stability of CLCuMuV-Raj and its spread toward the Sindh province where previously Cotton leaf curl Kokhran virus (CLCuKoV) and Cotton leaf curl Shahdadpur virus (CLCuShV) have been reported. The findings of the current study demand future research on CLCuD complex to explore the possible reasons for prevalence in the field and how the virus-host-vector compatible interaction can be broken to develop resistant cultivars.

Comparative analysis, diversification, and functional validation of plant nucleotide-binding site domain genes.

Nucleotide-binding site (NBS) domain genes are one of the superfamily of resistance genes involved in plant responses to pathogens. The current study identified 12,820 NBS-domain-containing genes across 34 species covering from mosses to monocots and dicots. These identified genes are classified into 168 classes with several novel domain architecture patterns encompassing significant diversity among plant species. Several classical (NBS, NBS-LRR, TIR-NBS, TIR-NBS-LRR, etc.) and species-specific structural patterns (TIR-NBS-TIR-Cupin_1-Cupin_1, TIR-NBS-Prenyltransf, Sugar_tr-NBS etc.) were discovered. We observed 603 orthogroups (OGs) with some core (most common orthogroups; OG0, OG1, OG2, etc.) and unique (highly specific to species; OG80, OG82, etc.) OGs with tandem duplications. The expression profiling presented the putative upregulation of OG2, OG6, and OG15 in different tissues under various biotic and abiotic stresses in susceptible and tolerant plants to cotton leaf curl disease (CLCuD). The genetic variation between susceptible (Coker 312) and tolerant (Mac7) Gossypium hirsutum accessions identified several unique variants in NBS genes of Mac7 (6583 variants) and Coker312 (5173 variants). The protein-ligand and proteins-protein interaction showed a strong interaction of some putative NBS proteins with ADP/ATP and different core proteins of the cotton leaf curl disease virus. The silencing of GaNBS (OG2) in resistant cotton through virus-induced gene silencing (VIGS) demonstrated its putative role in virus tittering. The presented study will be further helpful in understanding the plant adaptation mechanism.

Efficient A·T-to-C·G Base Editing via Adenine Transversion Editors.

Generating A-to-C transversions to correct defective alleles or introduce novel alleles has posed significant challenges. However, two recent studies focusing on adenine transversions have achieved successful A-to-C transversions in mouse embryos and plant cell. These remarkable accomplishments notably broaden the range of base editing and their applications both in fundamental research and in therapeutics.

Einkorn genomics reveals ancient roots of domesticated wheat.

Wheat is an important staple food crop that underwent complex genome duplications. During domestication, genetic changes occurred, improving modern wheat, but understanding its phylogenetic history has been lacking. Mahmood and Mansoor discuss a recent publication demonstrating the evolutionary history of domesticated wheat (Triticum monococcum), providing opportunities for advancements in cereal improvement.

PASTE: a high-throughput method for large DNA insertions.

Prime editing (PE) enables precise genome editing at targeted locus without inducing double-stranded breaks (DSBs). Despite its precision, PE lacks the tendency to integrate large DNA fragments into the genome. Recently, Yarnall et al. reported clustered regularly interspaced palindromic repeats (CRISPR)/Cas9 and an integrase-based system that conducts targeted integration of large DNA sequences (~36 kb) into the genome more efficiently.

Omics-driven exploration and mining of key functional genes for the improvement of food and fiber crops.

The deployment of omics technologies has obtained an incredible boost over the past few decades with the advances in next-generation sequencing (NGS) technologies, innovative bioinformatics tools, and the deluge of available biological information. The major omics technologies in the limelight are genomics, transcriptomics, proteomics, metabolomics, and phenomics. These biotechnological advances have modernized crop breeding and opened new horizons for developing crop varieties with improved traits. The genomes of several crop species are sequenced, and a huge number of genes associated with crucial economic traits have been identified. These identified genes not only provide insights into the understanding of regulatory mechanisms of crop traits but also decipher practical grounds to assist in the molecular breeding of crops. This review discusses the potential of omics technologies for the acquisition of biological information and mining of the genes associated with important agronomic traits in important food and fiber crops, such as wheat, rice, maize, potato, tomato, cassava, and cotton. Different functional genomics approaches for the validation of these important genes are also highlighted. Furthermore, a list of genes discovered by employing omics approaches is being represented as potential targets for genetic modifications by the latest genome engineering methods for the development of climate-resilient crops that would in turn provide great impetus to secure global food security.

Plant Virus-Derived Vectors for Plant Genome Engineering.

Advances in genome engineering (GE) tools based on sequence-specific programmable nucleases have revolutionized precise genome editing in plants. However, only the traditional approaches are used to deliver these GE reagents, which mostly rely on Agrobacterium-mediated transformation or particle bombardment. These techniques have been successfully used for the past decades for the genetic engineering of plants with some limitations relating to lengthy time-taking protocols and transgenes integration-related regulatory concerns. Nevertheless, in the era of climate change, we require certain faster protocols for developing climate-smart resilient crops through GE to deal with global food security. Therefore, some alternative approaches are needed to robustly deliver the GE reagents. In this case, the plant viral vectors could be an excellent option for the delivery of GE reagents because they are efficient, effective, and precise. Additionally, these are autonomously replicating and considered as natural specialists for transient delivery. In the present review, we have discussed the potential use of these plant viral vectors for the efficient delivery of GE reagents. We have further described the different plant viral vectors, such as DNA and RNA viruses, which have been used as efficient gene targeting systems in model plants, and in other important crops including potato, tomato, wheat, and rice. The achievements gained so far in the use of viral vectors as a carrier for GE reagent delivery are depicted along with the benefits and limitations of each viral vector. Moreover, recent advances have been explored in employing viral vectors for GE and adapting this technology for future research.

qPCR Assay as a Tool for Examining Cotton Resistance to the Virus Complex Causing CLCuD: Yield Loss Inversely Correlates with Betasatellite, Not Virus, DNA Titer.

Cotton leaf curl disease (CLCuD) is a significant constraint to the economies of Pakistan and India. The disease is caused by different begomoviruses (genus Begomovirus, family Geminiviridae) in association with a disease-specific betasatellite. However, another satellite-like molecule, alphasatellite, is occasionally found associated with this disease complex. A quantitative real-time PCR assay for the virus/satellite components causing CLCuD was used to investigate the performance of selected cotton varieties in the 2014-2015 National Coordinated Varietal Trials (NCVT) in Pakistan. The DNA levels of virus and satellites in cotton plants were determined for five cotton varieties across three geographic locations and compared with seed cotton yield (SCY) as a measure of the plant performance. The highest virus titer was detected in B-10 (0.972 ng·µg-1) from Vehari and the lowest in B-3 (0.006 ng·µg-1) from Faisalabad. Likewise, the highest alphasatellite titer was found in B-1 (0.055 ng·µg-1) from Vehari and the lowest in B-1 and B-2 (0.001 ng·µg-1) from Faisalabad. The highest betasatellite titer was found in B-23 (1.156 ng·µg-1) from Faisalabad and the lowest in B-12 (0.072 ng·µg-1) from Multan. Virus/satellite DNA levels, symptoms, and SCY were found to be highly variable between the varieties and between the locations. Nevertheless, statistical analysis of the results suggested that betasatellite DNA levels, rather than virus or alphasatellite DNA levels, were the important variable in plant performance, having an inverse relationship with SCY (-0.447). This quantitative assay will be useful in breeding programs for development of virus resistant plants and varietal trials, such as the NCVT, to select suitable varieties of cotton with mild (preferably no) symptoms and low (preferably no) virus/satellite. At present, no such molecular techniques are used in resistance breeding programs or varietal trials in Pakistan.

BioClay: next-generation crop protection strategy.

Whitefly and the viruses they transmit pose a serious threat to crops globally. Recently, Jain et al. showed that BioClay-mediated double-stranded RNA (dsRNA) spray provides an eco-friendly approach to controlling whitefly. This 'transgene-free next-generation' insect-specific crop protection strategy may help to reduce the use of chemical pesticides for controlling whitefly.

Dominance of Asia II 1 species of Bemisia tabaci in Pakistan and beyond.

Globally, Whitefly (Bemisia tabaci) is one of the most important insect pests of crops that causes huge economical losses. The current study was designed to exclusively screen the B. tabaci species in the cotton field of Pakistan during 2017-2020 and have to conduct comparative analysis of B. tabaci species in Asia where Asia II 1 has been reported. A total of 5142 B. tabaci sequences of mitochondrial cytochrome oxidase 1 (mtCO1) from Asian countries were analyzed to determine the species and their distribution in the region. Our analysis over time and space showed that Asia II 1 has gradually dominated over Asia 1 in Punjab Province and over both Asia 1 and MEAM1 in Sindh Province. Asia has been divided into three regions i.e., South Asia (2524 sequences), Southeast Asia (757 sequences) and East Asia (1569 sequences) and dominance of different species of B. tabaci has been determined by calculating the relative percentage of each species. Interestingly, Asia II 1 has been found dominant in the neighboring region (northern zone) of India and also being dominant in its central zone. The dominance of Asia II 1 in Pakistan and northern India explains whitefly epidemic being reported in recent years.

Genome wide study of cysteine rich receptor like proteins in Gossypium sp.

Cysteine-rich receptor-like-kinases (CRKs), a transmembrane subfamily of receptor-like kinase, play crucial roles in plant adaptation. As such cotton is the major source of fiber for the textile industry, but environmental stresses are limiting its growth and production. Here, we have performed a deep computational analysis of CRKs in five Gossypium species, including G. arboreum (60 genes), G. raimondii (74 genes), G. herbaceum (65 genes), G. hirsutum (118 genes), and G. barbadense (120 genes). All identified CRKs were classified into 11 major classes and 43 subclasses with the finding of several novel CRK-associated domains including ALMT, FUSC_2, Cript, FYVE, and Pkinase. Of these, DUF26_DUF26_Pkinase_Tyr was common and had elevated expression under different biotic and abiotic stresses. Moreover, the 35 land plants comparison identified several new CRKs domain-architectures. Likewise, several SNPs and InDels were observed in CLCuD resistant G. hirsutum. The miRNA target side prediction and their expression profiling in different tissues predicted miR172 as a major CRK regulating miR. The expression profiling of CRKs identified multiple clusters with co-expression under certain stress conditions. The expression analysis under CLCuD highlighted the role of GhCRK057, GhCRK059, GhCRK058, and GhCRK081 in resistant accession. Overall, these results provided primary data for future potential functional analysis as well as a reference study for other agronomically important crops.

Smart breeding approaches in post-genomics era for developing climate-resilient food crops.

Improving the crop traits is highly required for the development of superior crop varieties to deal with climate change and the associated abiotic and biotic stress challenges. Climate change-driven global warming can trigger higher insect pest pressures and plant diseases thus affecting crop production sternly. The traits controlling genes for stress or disease tolerance are economically imperative in crop plants. In this scenario, the extensive exploration of available wild, resistant or susceptible germplasms and unraveling the genetic diversity remains vital for breeding programs. The dawn of next-generation sequencing technologies and omics approaches has accelerated plant breeding by providing the genome sequences and transcriptomes of several plants. The availability of decoded plant genomes offers an opportunity at a glance to identify candidate genes, quantitative trait loci (QTLs), molecular markers, and genome-wide association studies that can potentially aid in high throughput marker-assisted breeding. In recent years genomics is coupled with marker-assisted breeding to unravel the mechanisms to harness better better crop yield and quality. In this review, we discuss the aspects of marker-assisted breeding and recent perspectives of breeding approaches in the era of genomics, bioinformatics, high-tech phonemics, genome editing, and new plant breeding technologies for crop improvement. In nutshell, the smart breeding toolkit in the post-genomics era can steadily help in developing climate-smart future food crops.

Geminiviruses also encode small proteins with specific functions.

Geminiviruses are small DNA viruses with single-stranded circular genomes that encode for few known proteins. Recently, Gong et al. reconsidered their protein identification criteria and identified open reading frames that encode small proteins with specific cellular localizations and virulence functions. Their data warrants exploration of such small proteins in other geminiviruses.