Recent advances in artificial intelligence, mechanistic models, and speed breeding offer exciting opportunities for precise and accelerated genomics-assisted breeding.

Given the challenges of population growth and climate change, there is an urgent need to expedite the development of high-yielding stress-tolerant crop cultivars. While traditional breeding methods have been instrumental in ensuring global food security, their efficiency, precision, and labour intensiveness have become increasingly inadequate to address present and future challenges. Fortunately, recent advances in high-throughput phenomics and genomics-assisted breeding (GAB) provide a promising platform for enhancing crop cultivars with greater efficiency. However, several obstacles must be overcome to optimize the use of these techniques in crop improvement, such as the complexity of phenotypic analysis of big image data. In addition, the prevalent use of linear models in genome-wide association studies (GWAS) and genomic selection (GS) fails to capture the nonlinear interactions of complex traits, limiting their applicability for GAB and impeding crop improvement. Recent advances in artificial intelligence (AI) techniques have opened doors to nonlinear modelling approaches in crop breeding, enabling the capture of nonlinear and epistatic interactions in GWAS and GS and thus making this variation available for GAB. While statistical and software challenges persist in AI-based models, they are expected to be resolved soon. Furthermore, recent advances in speed breeding have significantly reduced the time (3-5-fold) required for conventional breeding. Thus, integrating speed breeding with AI and GAB could improve crop cultivar development within a considerably shorter timeframe while ensuring greater accuracy and efficiency. In conclusion, this integrated approach could revolutionize crop breeding paradigms and safeguard food production in the face of population growth and climate change.

Chemical mutagenesis: role in breeding and biofortification of lentil (Lens culinaris Medik) mutant lines.

BACKGROUND: Induced mutagenesis is a quick and effective breeding strategy to enhance genetic variability, an important prerequisite for the genetic improvement of existing lentil cultivars. Lentil is an important cool season food legume with low productivity due to the low yielding potential of existing lentil cultivars. The present study aimed at increasing the yielding potential, resulted in the isolation of six high-yielding mutant lines with dense micronutrients. METHODS AND RESULTS: Two lentil varieties were treated with different doses of ethyl methanesulphonate, hydrazine hydrate, and sodium azide, followed by phenotypic selection for consecutive three generations. In the M2 generation, six high-yielding mutant lines with stable phenotypes were isolated. The results revealed a substantial increase in mean values for quantitative and physiological traits coupled with a manifold increase in the genotypic coefficient of variation (GCV), heritability (h2), and genetic advance (GA). Correlation analysis revealed that plant yield was significantly and positively influenced (P < 0.001) by fertile branches per plant, pods per plant, and seed weight. Principal component analysis revealed two principal components contributed 63.5 and 62.5% of the total variation in the varieties Pant L-639 and Pant L-406, respectively. CONCLUSION: The isolated high-yielding mutant lines with dense micronutrients that serve as rich genetic resources could be subjected to further breeding trials. After attaining yield stability, these might be registered and released as new improved lentil varieties.

Gamma rays and sodium azide induced variations in bio-physiological and agronomical traits in linseed (Linum usitatissimum L.).

Linseed is a valuable oilseed crop with huge therapeutic importance due to its high content of omega-3 fatty acids in the form of Alpha-linolenic acid (ALA). It is a self-pollinated crop with a low-yielding potential that restricts its improvement endeavors. To overcome low-yielding potential, individual and combination treatments of gamma rays and sodium azide were employed in widely grown linseed varieties. The results revealed a dose-dependent decline in seed germination, seedling height, pollen fertility, chlorophyll, and carotenoid contents and a dose-independent decline in carbonic anhydrase activity. Bio-physiological parameters decreased substantially in combination treatments compared to individual treatments of gamma rays and sodium azide. In contrast, lower doses of gamma rays, sodium azide, and their combinations effectively increased mean values of yield and yield-attributing traits in a few putative mutants. Such putative mutants represent a valuable genetic resource that could be used in future breeding programs for the genetic improvement of linseed and related medicinal plants.

Field assessment of yield and its contributing traits in cowpea treated with lower, intermediate, and higher doses of gamma rays and sodium azide.

Across the globe, plant breeders of different organizations are working in collaboration to bring preferred traits to crops of economic importance. Among the traits, "high yielding potential" is the most important as it is directly associated with food security and nutrition, one of the sustainable development goals. The Food and Agriculture Organization acknowledges plant breeders' role and efforts in achieving local and global food security and nutrition. Recognizing the importance of pulses and increasing pressure on food security, the United Nations General Assembly declared 2016 the "International year of Pulses" owing to their preferred traits such as climate change resilience, wide adaptability, low agriculture input, and protein- and nutrient-rich crops. Keeping all these developments in consideration, we initiated an induced mutagenesis program by treating cowpea (Vigna unguiculata L. Walp.) with different doses of gamma rays and sodium azide aiming to enhance the yielding potential of an otherwise outstanding variety viz., Gomati VU-89 and Pusa-578. We noticed a substantial increase in mean values of agronomic traits in putative mutants raised from seeds treated with lower and intermediate doses of mutagens. Statistical analysis such as correlation, path, hierarchical clustering analysis (HCA), and principal component analysis (PCA) were used to assess the difference between mutagenized and control populations. A significant and positive correlation of yield with yield-attributing traits was recorded. However, among all the yield attributing traits, seeds per pod (SPP) depicted the maximum direct impact upon yield, and therefore, working on this trait may yield better results. A widely used PCA revealed 40.46% and 33.47% of the total variation for var. Gomati VU-89 and var. Pusa-578, respectively. Cluster analysis clustered treated and control populations into separate clusters with variable cluster sizes. Cluster V in the variety Gomati VU-89 and cluster V and VI in the variety Pusa 578 comprised of putative mutants were higher yielding and hence could be recommended for selection in future breeding programs. We expect to release such mutant lines for farmer cultivation in Northern parts of India depending on the performance of such high-yielding mutant lines at multilocations.

Assessment of Bio-physiological damages and cytological aberrations in cowpea varieties treated with gamma rays and sodium azide.

The assessment of mutagen induced biological damage forms an important study in determining the mutagenic potency and genotypic sensitivity, a vital aspect in mutation breeding programs. A prior assessment of lethal dose (LD50), mutagen induced biological damage (alterations in bio-physiological traits and frequency of cytological aberrations) is a prerequisite for determining an optimum mutagen dose in a successful mutation breeding experiment. Therefore, in a multi-year project of mutation breeding, two widely cultivated varieties of cowpea viz., Gomati VU-89 and Pusa-578, were treated with gamma (γ) rays and sodium azide (SA) doses. The results reflected a proportionate increase in bio-physiological damages with the increase in mutagenic doses and caused a substantial reduction in mean seed germination and seedling height. Different cytological aberrations such as cytomixis, univalents, chromosome stickiness, precocious separation, unequal separation, bridges, laggards, disturbed polarity, dyads, triads, and polyads were observed in both varieties. All the mutagen doses induced a broader spectrum of cytological aberrations with varying frequencies.

Comparative effects of caffeine and lead nitrate on the bio-physiological and yield associated traits of lentil (Lens culinaris Medik.).

Lentil belonging to the fabaceae family is a proteinaceous cool-season legume consumed across the world. However, lentil is low yielding with a narrow genetic base compared to other grain legumes such as chickpea, faba bean, and cowpea. In the present study, we intended to investigate the effect of two different mutagens viz., caffeine and lead nitrate on the bio-physiological and agronomical traits of lentil. Unlike other mutagens like ethyl methanesulphonate, sodium azide, and hydrazine hydrates very little is known about the mutagenic potency of caffeine and lead nitrate. The results revealed contrasting effects as lower doses of caffeine-induced a substantial increase in mean values of physiological and agronomical traits whereas both lower and higher doses of lead nitrate negatively impacted the agronomical traits of lentil. Among the mutagen doses, 0.1% caffeine was most efficient in inducing a substantial increase in mean values of bio-physiological and quantitative traits. The present study also revealed the successful conduct of induced mutagenesis in lentil and present a protocol that could be followed in future breeding programs aimed at increasing the yielding potential of legumes.

Chromium Toxicity in Plants: Signaling, Mitigation, and Future Perspectives.

Plants are very often confronted by different heavy metal (HM) stressors that adversely impair their growth and productivity. Among HMs, chromium (Cr) is one of the most prevalent toxic trace metals found in agricultural soils because of anthropogenic activities, lack of efficient treatment, and unregulated disposal. It has a huge detrimental impact on the physiological, biochemical, and molecular traits of crops, in addition to being carcinogenic to humans. In soil, Cr exists in different forms, including Cr (III) "trivalent" and Cr (VI) "hexavalent", but the most pervasive and severely hazardous form to the biota is Cr (VI). Despite extensive research on the effects of Cr stress, the exact molecular mechanisms of Cr sensing, uptake, translocation, phytotoxicity, transcript processing, translation, post-translational protein modifications, as well as plant defensive responses are still largely unknown. Even though plants lack a Cr transporter system, it is efficiently accumulated and transported by other essential ion transporters, hence posing a serious challenge to the development of Cr-tolerant cultivars. In this review, we discuss Cr toxicity in plants, signaling perception, and transduction. Further, we highlight various mitigation processes for Cr toxicity in plants, such as microbial, chemical, and nano-based priming. We also discuss the biotechnological advancements in mitigating Cr toxicity in plants using plant and microbiome engineering approaches. Additionally, we also highlight the role of molecular breeding in mitigating Cr toxicity in sustainable agriculture. Finally, some conclusions are drawn along with potential directions for future research in order to better comprehend Cr signaling pathways and its mitigation in sustainable agriculture.

Harnessing the potential of bulk segregant analysis sequencing and its related approaches in crop breeding.

Most plant traits are governed by polygenes including both major and minor genes. Linkage mapping and positional cloning have contributed greatly to mapping genomic loci controlling important traits in crop species. However, they are low-throughput, time-consuming, and have low resolution due to which their efficiency in crop breeding is reduced. In this regard, the bulk segregant analysis sequencing (BSA-seq) and its related approaches, viz., quantitative trait locus (QTL)-seq, bulk segregant RNA-Seq (BSR)-seq, and MutMap, have emerged as efficient methods to identify the genomic loci/QTLs controlling specific traits at high resolution, accuracy, reduced time span, and in a high-throughput manner. These approaches combine BSA with next-generation sequencing (NGS) and enable the rapid identification of genetic loci for qualitative and quantitative assessments. Many previous studies have shown the successful identification of the genetic loci for different plant traits using BSA-seq and its related approaches, as discussed in the text with details. However, the efficiency and accuracy of the BSA-seq depend upon factors like sequencing depth and coverage, which enhance the sequencing cost. Recently, the rapid reduction in the cost of NGS together with the expected cost reduction of third-generation sequencing in the future has further increased the accuracy and commercial applicability of these approaches in crop improvement programs. This review article provides an overview of BSA-seq and its related approaches in crop breeding together with their merits and challenges in trait mapping.

Conventional and new-breeding technologies for improving disease resistance in lentil (Lens culinaris Medik).

Lentil, an important cool season food legume, is a rich source of easily digestible protein, folic acid, bio-available iron, and zinc nutrients. Lentil grows mainly as a sole crop in the winter after harvesting rice in South Asia. However, the annual productivity is low due to its slow growth during the early phase, competitive weed infestation, and disease outbreaks during the crop growth period. Disease resistance breeding has been practiced for a long time to enhance resistance to various diseases. Often the sources of resistance are available in wild crop relatives. Thus, wide hybridization and the ovule rescue technique have helped to introgress the resistance trait into cultivated lentils. Besides hybridization, induced mutagenesis contributed immensely in creating variability for disease tolerance, and several disease-resistant mutant lines have been developed. However, to overcome the limitations of traditional breeding approaches, advancement in molecular marker technologies, and genomics has helped to develop disease-resistant and climate-resilient lentil varieties with more precision and efficiency. This review describes types of diseases, disease screening methods, the role of conventional and new breeding technologies in alleviating disease-incurred damage and progress toward making lentil varieties more resilient to disease outbreaks under the shadow of climate change.

Gamma Rays and Sodium Azide Induced Genetic Variability in High-Yielding and Biofortified Mutant Lines in Cowpea [Vigna unguiculata (L.) Walp.].

With the twin pressures of high population growth and extreme weather events, developing countries are the worst hit in meeting the food demands of their people, with millions unable to access adequate and nutritionally balanced food. Crop production must be increased by 70% to keep up with the food demands of a rapidly growing population, which is expected to rise to 9.6 billion by 2050. Legumes are ideal food crops to increase agricultural productivity and achieve sustainable development goals. Cowpea, a warm-season grain legume, is often categorized as a neglected crop with immense scope for genetic improvement through proper breeding strategies. A multi-year field experiment of induced mutagenesis was conducted to increase seed yield and genetic variability in the agro-economic traits of two cowpea varieties treated with different doses of gamma (γ) rays and sodium azide (SA). The study was also aimed to optimize different doses of γ rays and SA employed individually and in combinations. Quantitative trait analysis revealed a maximum increase in seed yield from M2 to M3 generation. Among the 10 quantitative traits studied, seeds per pod and seed weight positively correlated with a major direct impact on yield. An extensive phenotypic selection cycle from M2-M4 generations resulted in isolating new high-yielding and nutrient-dense mutant lines. Such high-yielding biofortified mutant lines with enhanced genetic variability could serve as a donor of elite genes and represent a valuable genetic resource for improving low-yielding warm-season grain legumes.

Comparative Mutagenic Effectiveness and Efficiency of Gamma Rays and Sodium Azide in Inducing Chlorophyll and Morphological Mutants of Cowpea.

Mutagenic effectiveness and efficiency are the most important factors determining the success of mutation breeding, a coherent tool for quickly enhancing genetic diversity in crops. However, conclusive evidence of using an effective and efficient dose of gamma (γ) rays and sodium azide (SA) for genetic improvement is scant. The present study assesses genetic diversity in M2 mutants of cowpea and evaluates mutagenic effectiveness and efficiency of the single and combination doses of γ rays and SA. In M0 generation, 7200 M1 seeds obtained by SA treatment (0.01-0.1%) and γ irradiation (100-1000 Gy) at a dose rate of 11.58 Gy/min were sown to raise M1 generation. A total of 57,620 M2 seeds were generated from the M1 generation of two varieties-Gomati VU-89 and Pusa-578, from which 47,650 seeds germinated. Moreover, plants (38,749) that survived were screened for chlorophyll and morphological mutations. Among the mutagens, SA followed by γ rays + SA and γ rays was most effective in inducing higher frequency and a broader spectrum of chlorophyll mutants. A wide range of morphological mutants affecting every growth stage was recorded with the highest frequency in 400 Gy γ rays + 0.04% SA treatment. These morphological mutants with desirable agronomic traits represent a valuable genetic resource for future breeding programs. This study revealed the potency of γ rays and SA in increasing genetic diversity and demonstrated the successful conduct of induced mutagenesis in the cowpea.

Phenotypic diversity in mutagenized population of urdbean (Vigna mungo (L.) Hepper).

The present study was conducted to assess the extent of induced genetic variability and to determine the inheritance pattern of various yield contributing phenotypic traits in M2 and M3 generations of urdbean following mutagenesis with single and combination treatments of gamma rays and ethyl methanesulphonate (EMS). The mean number of seeds per pod and 100-seed weight increased in all the mutagenic treatments in both the varieties with few exceptions in M2 generation. Mean pod length although increased considerably, however it did not differ significantly in most of the mutagenic treatments. In M3 generation, 0.2% EMS and 300 Gy γ rays+0.2% EMS treatments induced maximum increase in mean pod length, seeds per pod and 100-seed weight in both the varieties. Genetic parameters showed manifold increase in most of the mutagenic treatments and varied from trait to trait. Increased genetic variability for economically important traits in the selected mutant lines has successfully contributed in diversifying the accessible genetic base which could be exploited for subsequent improvement of urdbean through phenotypic selection.

Mechanisms of Genome Maintenance in Plants: Playing It Safe With Breaks and Bumps.

Maintenance of genomic integrity is critical for the perpetuation of all forms of life including humans. Living organisms are constantly exposed to stress from internal metabolic processes and external environmental sources causing damage to the DNA, thereby promoting genomic instability. To counter the deleterious effects of genomic instability, organisms have evolved general and specific DNA damage repair (DDR) pathways that act either independently or mutually to repair the DNA damage. The mechanisms by which various DNA repair pathways are activated have been fairly investigated in model organisms including bacteria, fungi, and mammals; however, very little is known regarding how plants sense and repair DNA damage. Plants being sessile are innately exposed to a wide range of DNA-damaging agents both from biotic and abiotic sources such as ultraviolet rays or metabolic by-products. To escape their harmful effects, plants also harbor highly conserved DDR pathways that share several components with the DDR machinery of other organisms. Maintenance of genomic integrity is key for plant survival due to lack of reserve germline as the derivation of the new plant occurs from the meristem. Untowardly, the accumulation of mutations in the meristem will result in a wide range of genetic abnormalities in new plants affecting plant growth development and crop yield. In this review, we will discuss various DNA repair pathways in plants and describe how the deficiency of each repair pathway affects plant growth and development.

Characterization of Induced High Yielding Cowpea Mutant Lines Using Physiological, Biochemical and Molecular Markers.

Cowpea, Vigna unguiculata (L.) Walp. is an important grain legume grown in the dry agro-ecologies of the tropics with considerably low yield due to lack of improved varieties, aggravated by prevalent narrow genetic base. Thus, induced mutagenesis was employed using sodium azide and gamma rays to increase genetic variability in cowpea genotypes that resulted in isolation of eleven high yielding mutant lines at the M4 generation from the genetic background of cowpea varieties Gomati VU-89 and Pusa-578. In order to analyze the induced genetic divergence among the mutant lines and parent genotypes, biochemical and molecular characterization was carried out with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), simple sequence repeat (SSR) and CAAT box derived polymorphism (CBDP) markers. Activity of nitrate reductase (NR) and content of chlorophyll, carotenoid, protein and mineral were found to be significantly high in the selected mutant lines compared to their respective parent genotypes. SDS-PAGE profile of seed proteins generated 54 and 28 polymorphic bands and a total polymorphism of 62.06 and 41.17% in Gomati VU-89 and Pusa-578, respectively. SSR primers amplified a total of 16 and 24 polymorphic bands with an average polymorphism of 20.69 and 50.74% in Gomati VU-89 and Pusa-578, respectively. CBDP markers, used for the first time in mutagenized population, generated 175 bands with 77 bands being polymorphic in Gomati VU-89 and 121 bands with 59 bands being polymorphic in Pusa-578. Physiological, biochemical and molecular profiling of the selected promising mutants lines showed that Gomati VU-89-G and Pusa-578-C are genetically most diverged high yielding genotypes with significant increase in protein and micronutrient content, therefore, could be recommended for further research considerations. Thus, the favorable combination of genes induced in the novel cowpea mutants selected through the present study are valuable to correspond farmers requirements for new improved cultivars (direct or hybrids).

Morphological characterization of gamma rays induced multipodding mutant (mp) in lentil cultivar Pant L 406.

PURPOSE: Lentil (Lens culinaris Medik.), being a self-pollinated crop with narrow genetic base, is an important target crop for mutation breeding experiments. The purpose of the investigation was to create, select and characterize unique mutations in inflorescence traits that have strong impact on lentil yield and yield stability. MATERIALS AND METHODS: Healthy and uniform seeds (moisture 11.0%) of Lens culinaris Medik. cultivar Pant L 406 were irradiated with 100, 200, 300 and 400 Gy of gamma rays. The mutagenized populations were maintained up to mutant generation third (M3) to screen for stable mutations in the inflorescence architecture of the lentil. The selected mutant mp 'Multipodding' trait, i.e. multiple pods per peduncle, was morphologically characterized and quantified in subsequent mutant generation fourth (M4). RESULTS: The morphological characterization of the 'multipodding' mutant (mp) revealed substantial morphological mutations were induced by the treatment of gamma rays. The estimation of yield per plant (g) between the mutant (mp) and parent cultivar Pant L 406 showed non-significant variation due to significant reduction in seed weight. CONCLUSIONS: The novel 'multipodding' (mp) mutant induced in the present study can play a key role in understanding the genetic network controlling legume inflorescence architecture and in genomics-assisted breeding for development of elite lentil cultivars.

Induced mutation analysis with biochemical and molecular characterization of high yielding lentil mutant lines.

Induced mutagenesis generates macromolecular variations which ultimately alters the bio-physiological and morphological nature of the crop genotypes. In the present study, molecular characterization of six high yielding lentil mutant lines, developed from hydrazine hydrates (HZ) and gamma rays mutagenesis, was carried out with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and random amplified polymorphic DNA (RAPD) markers. Activity of nitrate reductase (NR) and content of chlorophyll and carotenoid were found to be significantly high in the mutant lines. Protein and mineral (Fe, Zn & Cu) contents were also increased considerably in the mutant lines compared to their respective parent genotypes. SDS-PAGE profile of seed storage proteins showed 35 unique bands with 97.14% polymorphism. Genetic divergence analysis generated total 41 reproducible RAPD bands with average calculated polymorphic percentage of 63.06%. Among the primers, OPA-10 showed the highest polymorphism with significant PIC value. Genetic divergent analysis revealed that genome of cultivar DPL 62 mutated relatively more than the cultivar Pant L 406 due to the mutagen treatments, while DPL 62-B and Pant L406-A were the most divergent mutants induced in the present study. Biochemical and molecular profile of the induced mutant lines facilitates a basis for future conservation and utilization strategies to widen the genetic base of the current lentil breeding population.